Nothing
R/dbtool.R
In PGA: An package for identification of novel peptides by customized database derived from RNA-Seq
Defines functions
getNovelTx
.gen_splicmatrix
PrepareAnnotationRefseq2
PrepareAnnotationEnsembl2
OutputVarproseq2
OutputNovelJun2
Outputaberrant2
Tab2Range
Documented in
PrepareAnnotationEnsembl2
PrepareAnnotationRefseq2
## This function requires the Tab format file from Hisat2. Read Tab file contain junctions into a GRanges object.
Tab2Range <- function(tabfile)
{
options(stringsAsFactors=FALSE)
jun <- read.table(tabfile, sep='\t', header=F, stringsAsFactors=F)
#replace the strand "." with "+" and "-".
tmp<-jun[jun$V4==".",]
tmp_forward<-tmp
tmp_reverse<-tmp
tmp_forward$V4<-"+"
tmp_reverse$V4<-"-"
jun<-rbind(jun[jun$V4!=".",],tmp_forward,tmp_reverse)
part1_sta <- as.numeric(jun[,'V2'])+1-59
part1_end <- as.numeric(jun[,'V2'])+1
part2_sta <- as.numeric(jun[,'V3'])+1
part2_end <- as.numeric(jun[,'V3'])+1+59
junction <- data.frame(chr=jun[, 'V1'], id="NULL", start=jun[, 'V2'],
end=jun[,'V3'], cov=255, strand=jun$V4, part1_len=0,
part2_len=0, part1_sta, part1_end, part2_sta, part2_end)
if('chrM' %in% junction$chr) junction <- junction[-which(junction$chr=='chrM'), ]
if('MT' %in% junction$chr) junction <- junction[-which(junction$chr=='MT'), ]
junRange <- GRanges(seqnames=junction$chr, ranges=IRanges(start=junction$part1_end,
end=junction$part2_sta), strand=junction$strand, id=junction$id,
cov=junction$cov, part1_len=junction$part1_len, part2_len=junction$part2_len,
part1_sta=junction$part1_sta, part1_end=junction$part1_end,
part2_sta=junction$part2_sta, part2_end=junction$part2_end)
}
Outputaberrant2 <- function(positiontab, outfa, outmtab, coding, proteinseq,
ids, RPKM=NULL, ...){
options(stringsAsFactors=FALSE)
idx <- grep(',', positiontab[, 'varbase'], fixed=TRUE)
if(length(idx) > 0) {
muti <- positiontab[idx, ]
muti_new <- c()
for(i in 1:dim(muti)[1]){
tmp <- cbind(muti[i, 1:8],
unlist(strsplit(muti[i, 'varbase'], ',', fixed=TRUE)),
muti[i, 10])
muti_new <- rbind(muti_new, tmp)
}
colnames(muti_new) <- colnames(positiontab)
positiontab <- positiontab[-idx, ]
positiontab <- rbind(positiontab, muti_new)
}
mtable <- merge(positiontab, coding, by.x='txid', by.y='tx_id', all.x=TRUE,
stringsAsFactors = FALSE)
#mtable <- cbind(positiontab,codingseq)
mtab_plus <- subset(mtable, strand == '+')
mtab_minus <- subset(mtable, strand == '-')
str_sub(mtab_plus[, 'coding'], mtab_plus[, 'pincoding'],
nchar(mtab_plus[, 'refbase']) + mtab_plus[, 'pincoding'] - 1) <-
mtab_plus[, 'varbase']
str_sub(mtab_minus[, 'coding'], mtab_minus[, 'pincoding'],
mtab_minus[, 'pincoding'] + nchar(mtab_minus[, 'refbase']) - 1) <-
as.character(reverseComplement(DNAStringSet(mtab_minus[, 'varbase'])))
total <- rbind(mtab_plus,mtab_minus)
total<-subset(total,!is.na(coding),) # added in 20161027, some protein ids didn't have the corresponding coding sequence. Therefore, the DNAStringSet would get the error.
label <- unlist(lapply(total[,'coding'], function(x){
if(grepl ('N',x,fixed=TRUE)){
paste('with N in pos ', paste(as.character(
gregexpr('N', x, fixed=TRUE)[[1]]), collapse=' '),
sep='')
}else ""}))
total[,'coding'] <- gsub('N', 'A', total[, 'coding'], fixed=TRUE)
aa <- suppressWarnings(as.character(translate(DNAStringSet(total[, 'coding']))))
total <- cbind(total, aa, label)
pep <- apply(total, 1, function(x) unlist(strsplit(x['aa'], '\\*'))[1])
total <- cbind(total, pep)
#########remove the entry identical to normal sequence
plist <- unique(total[, 'proname'])
proteinseq <- subset(proteinseq, pro_name %in% plist)
mseq <- merge(total,proteinseq, by.x='proname', by.y='pro_name',
all=FALSE, stringsAsFactors = FALSE)
#index <- apply(mseq, 1, function(x) grep(x['pep'],x['peptide'],fixed=T))
#grep(mseq[4,'pep'],mseq[4,'peptide'])
index <-c()
for(i in 1:dim(mseq)[1]){
index <- c(index, ifelse(grep(mseq[i, 'pep'],
mseq[i, 'peptide'], fixed=TRUE) == 1, i, '' ))
#print(i)
}
if(length(index) > 0) mseq_r <- mseq[-index, ] else mseq_r <- mseq
mseq_f <- merge(mseq_r, ids, by.x='pro_name', by.y='pro_name',
all=FALSE, stringsAsFactors = FALSE)
#vartable<-cbind(Index=paste("SNV",rownames(vartable),sep=""),vartable)
mseq_f<-cbind(Index=paste("IDL",rownames(mseq_f),sep=""),mseq_f)
mseq_o<-subset(mseq_f,select=c(Index,txid,genename,txname,proname,chr,
strand,pos,refbase,varbase,pincoding,label))
write.table(mseq_o, file=outmtab, sep='\t', quote=FALSE, row.names=FALSE)
subset(mseq_f,select=c(pro_name,proname,txid,genename,txname,chr,strand,
pos,refbase,varbase,pincoding,label,gene_name,
tx_name))
outformat <- paste('>',mseq_f[,"Index"],"|", mseq_f[, 'pro_name'], "|",
mseq_f[, 'pincoding'],
" ", mseq_f[, 'refbase'], '>', mseq_f[, 'varbase'], "|",
mseq_f[, 'tx_name'], "|", mseq_f[, 'gene_name'], "|",
mseq_f[, 'description'], "|", mseq_f[, 'label'], '\n',
mseq_f[, 'pep'], sep='')
write(outformat, file=outfa)
}
OutputNovelJun2 <- function(junction_type, genome, outfa,outmtab,
#outfile_c,
proteinseq, debug=FALSE, ...){
options(stringsAsFactors=FALSE)
#ids <- subset(ids,pro_name!='')
#trans <- transcripts(txdb)
#index <- which(values(trans)[['tx_name']] %in% ids[,'tx_name'])
#pro_trans <- trans[index]
novel_junc <- subset(junction_type, jun_type != 'known junction')
if(!length(grep('chr', novel_junc[, 'seqnames'], fixed=TRUE))>0) {
novel_junc[, 'seqnames'] <- paste('chr', novel_junc[, 'seqnames'],
sep='')#
idx <- which(novel_junc[, 'seqnames'] %in% seqnames(genome))
novel_junc <- novel_junc[idx, ]#
}
###remove abnormal junctions
idx_abn <- union(which(novel_junc[, 'start'] < 0),
which(novel_junc[, 'end'] < 0))
if(length(idx_abn > 0)) novel_junc <- novel_junc[-idx_abn, ]
junRange1 <- GRanges(seqnames=novel_junc$seqnames,
ranges=IRanges(start=novel_junc$part1_sta,
end=novel_junc$part1_end),
strand=novel_junc$strand,
junction_id=novel_junc$id)
junRange2 <- GRanges(seqnames=novel_junc$seqnames,
ranges=IRanges(start=novel_junc$part2_sta,
end=novel_junc$part2_end),
strand=novel_junc$strand,
junction_id=novel_junc$id)
#match1_protx <- findOverlaps(junRange1,pro_trans)
#match2_protx <- findOverlaps(junRange2,pro_trans)
#juntransRange1 <- junRange1[unique(queryHits(match1_protx))]
#juntransRange2 <- junRange2[unique(queryHits(match2_protx))]
#junseq1 <- getSeq(genome,'chr1',start=1000,end=2000,as.character=TRUE)
###already did reverseComplement
junseq1 <- getSeq(genome, junRange1)
junseq2 <- getSeq(genome, junRange2)
junseq_cat <- DNAStringSet(mapply(function(x, y, z)
ifelse(z == '+', paste(x, y, sep=''), paste(y, x, sep='')),
as.data.frame(junseq1)[, 1],
as.data.frame(junseq2)[, 1], as.character(strand(junRange1))))#
#debug
debug=T
if(debug){
debug_file<-data.frame(seqnames=novel_junc$seqnames, part1_sta=novel_junc$part1_sta,
part1_end=novel_junc$part1_end,part2_sta=novel_junc$part2_sta,
part1_end=novel_junc$part2_end,strand=novel_junc$strand, seq1=junseq1, seq2=junseq2, seqcat=junseq_cat)
write.table(debug_file,file="debug_file.tsv",quote = F, row.names=F)
}
#index_plus <- which(strand(junRange1) == '+')
#index_minus <- which(strand(junRange1) == '-')
#seqs_plus <- junseq_cat[index_plus]
#seqs_minus <- reverseComplement(junseq_cat[index_minus])
#seqs <- c(seqs_plus, seqs_minus)
#novel_junc_new <- rbind(novel_junc[index_plus, ],
# novel_junc[index_minus, ])
novel_junc_new <- novel_junc
seqs <- junseq_cat
##Remove sequences contains NNN
Nindx <- grep('N', seqs)
if(length(Nindx) > 0){
seqs <- seqs[-Nindx]
novel_junc_new <- novel_junc_new[-Nindx, ]
}
seqs_name <- paste(paste(novel_junc_new[, 'id'], '_',
novel_junc_new[, 'seqnames'], ':',
novel_junc_new[, 'start'], '-',
novel_junc_new[, 'end'],
sep=''), novel_junc_new[, 'cov'],sep='|')#
junpepcoding <- data.frame('pro_name'=seqs_name,
'coding'=as.data.frame(seqs)[, 1])
#save(junpepcoding, file=outfile_c)
peptides_r1 <- suppressWarnings(translate(seqs))
peptides_r2 <- suppressWarnings(translate(subseq(seqs, start=2)))
peptides_r3 <- suppressWarnings(translate(subseq(seqs, start=3)))
junpos_rna_p1 <- ifelse(novel_junc_new[, 'strand'] == '+',
as.numeric(novel_junc_new[, 'part1_len']),
as.numeric(novel_junc_new[, 'part2_len'])) #
junpos_rna_p2 <- ifelse(novel_junc_new[, 'strand'] == '+',
as.numeric(novel_junc_new[, 'part1_len'])+1,
as.numeric(novel_junc_new[, 'part2_len'])+1) #
junpos_r1_p1 <- ceiling(junpos_rna_p1/3)#
junpos_r1_p2 <- ceiling(junpos_rna_p2/3) #
junpos_r2_p1 <- ceiling((junpos_rna_p1-1)/3) #
junpos_r2_p2 <- ceiling((junpos_rna_p2-1)/3)
junpos_r3_p1 <- ceiling((junpos_rna_p1-2)/3) #
junpos_r3_p2 <- ceiling((junpos_rna_p2-2)/3)
rownum<-dim(novel_junc_new)[1]
outdf<-rbind(cbind(Index=paste("JUC",1:rownum,sep=""),
frame=rep("1",rownum),
junpos_p1=junpos_r1_p1,
junpos_p2=junpos_r1_p2,
novel_junc_new,
sequence=as.data.frame(peptides_r1)[, 1]),
cbind(Index=paste("JUC",(rownum+1):(2*rownum),sep=""),
frame=rep("2",rownum),
junpos_p1=junpos_r2_p1,
junpos_p2=junpos_r2_p2,
novel_junc_new,
sequence=as.data.frame(peptides_r2)[, 1]),
cbind(Index=paste("JUC",(2*rownum+1):(3*rownum),sep=""),
frame=rep("3",rownum),
junpos_p1=junpos_r3_p1,
junpos_p2=junpos_r3_p2,
novel_junc_new,
sequence=as.data.frame(peptides_r3)[, 1]))
### remove peptide contain stop codon
index_stop <- grep('*', outdf$sequence, fixed=TRUE)
if(length(index_stop) > 0){
outdf_rmstop <- outdf[-index_stop, ]
}else outdf_rmstop <- outdf
### check if any peptides can be found in the normal database, remove those
index_nor <- lapply(outdf_rmstop$sequence, function(x)
grep(x, proteinseq[, 'peptide'], fixed=TRUE))
index_nor <- which(unlist(lapply(index_nor, length)) > 0)
if(length(index_nor) > 0){
outdf_new <- outdf_rmstop[-index_nor, ]
}else outdf_new <- outdf_rmstop
write.table(outdf_new[,-29], file=outmtab, sep='\t', quote=FALSE,
row.names=FALSE)
tmp<-paste(">",outdf_new[,'Index'],"|",outdf_new[,'seqnames'],":",
outdf_new[,"start"],"-",outdf_new[,"end"],"|ORF",
outdf_new[,"frame"]," Junpos:",outdf_new[,"junpos_p1"],"-",
outdf_new[,"junpos_p2"],"|",outdf_new[,"strand"],"|",
outdf_new[,"tx_name_part1"],"|",outdf_new[,"tx_name_part2"],"|",
outdf_new[,"jun_type"],"\n",outdf_new[,"sequence"],sep="")
write(tmp, file=outfa)
}
OutputVarproseq2 <- function(vartable, proteinseq, outfa, outmtab, ids, lablersid=FALSE,
RPKM=NULL, ...){
options(stringsAsFactors=FALSE)
abnor_index<-which(vartable$aavar=="non-synonymous")
if(length(abnor_index)!=0){
vartable<-vartable[-abnor_index,]
}
abnor_index<-which(vartable$aapos==1 & vartable$aavar=="*")
if(length(abnor_index)!=0){
vartable<-vartable[-abnor_index,]
}
vartable<-cbind(Index=paste("SNV",rownames(vartable),sep=""),vartable)
write.table(vartable, file=outmtab, sep='\t', quote=FALSE, row.names=FALSE)
nonsy <- vartable[vartable[, 'vartype'] == "non-synonymous", ]
if(lablersid){
aavar2pro <- subset(nonsy, select=c(Index,genename, txname, proname,
aaref, aapos, aavar, rsid))
}else{
aavar2pro <- subset(nonsy, select=c(Index,genename, txname, proname,
aaref, aapos, aavar))
}
aavar2pro <- aavar2pro[aavar2pro[, 'aaref']!="*", ]
#aavar2pro <- aavar2pro[aavar2pro[, 'aavar']!="*", ]
aavar2pro <- unique(aavar2pro)
plist <- unique(aavar2pro[, 'proname'])
pep <- proteinseq[proteinseq[, 'pro_name'] %in% plist, ]
pep_var <- pep
pep_all<- c()
test <- c()
for(i in 1:dim(pep_var)[1]){
#print(i)
pvar <-subset(aavar2pro,aavar2pro[,'proname'] == pep_var[i,'pro_name'])
pvar <- pvar[order(as.numeric(pvar[, 'aapos'])), ]
for(j in 1:dim(pvar)[1]){
tmp_pep_var<-pep_var[i,]
substr(tmp_pep_var[1, 'peptide'], as.integer(pvar[j, 'aapos']),
as.integer(pvar[j, 'aapos'])) <- substr(pvar[j,'aavar'],1,1)
if(tmp_pep_var[1, 'peptide']!=pep[i, 'peptide']){
if(lablersid){
var_name<-ifelse(is.null(pvar[j,'rsid']),
paste(pvar[j,"aaref"],pvar[j,"aapos"],
pvar[j,"aavar"],sep=""),
paste(pvar[j,"rsid"],":",pvar[j,"aaref"],
pvar[j,"aapos"],pvar[j,"aavar"],
sep=""))
}else{
var_name <- paste(pvar[j,"aaref"],pvar[j,"aapos"],
pvar[j,"aavar"],sep="")}
pep_name <- cbind(Index=pvar[j,"Index"],tmp_pep_var[1,],
var_name=toString(var_name))
pep_all <- rbind(pep_all, pep_name)
}else{
test <- c(test,tmp_pep_var[1, 'pro_name'])
}
}
}
#save(pep_all,file="pep_all.Rdata")
ftab <- merge(pep_all, ids, by.x='pro_name', by.y='pro_name', all=FALSE,
stringsAsFactors = FALSE)
outformat <- apply(ftab, 1, function(x)
paste('>',x['Index'],"|", x['pro_name'], "|", x['var_name'], " ",
x['tx_name.x'], "|", x['gene_name'], "|", x['description'],
'\n', unlist(strsplit(x['peptide'], '\\*'))[1], sep=''))
write(outformat, file=outfa)
}
#' @title Prepare annotation from ENSEMBL
#' @description Prepare the annotation from ENSEMBL through biomaRt. This
#' function is modified from the function \code{\link{PrepareAnnotationEnsembl}} in \pkg{customProDB}.
#' @param mart See detail in function \code{\link{PrepareAnnotationEnsembl}}.
#' @param annotation_path See detail in function \code{\link{PrepareAnnotationEnsembl}}.
#' @param splice_matrix See detail in function \code{\link{PrepareAnnotationEnsembl}}.
#' @param dbsnp See detail in function \code{\link{PrepareAnnotationEnsembl}}.
#' @param transcript_ids See detail in function \code{\link{PrepareAnnotationEnsembl}}.
#' @param COSMIC See detail in function \code{\link{PrepareAnnotationEnsembl}}.
#' @param ... Additional arguments
#' @return Several .RData file containing annotations needed for following analysis.
#' @seealso \code{\link{PrepareAnnotationRefseq2}}.
#' @export
#' @examples
#' ensembl <- biomaRt::useMart("ENSEMBL_MART_ENSEMBL", dataset="hsapiens_gene_ensembl",
#' host="grch37.ensembl.org", path="/biomart/martservice",
#' archive=FALSE)
#'
#' annotation_path <- tempdir()
#' transcript_ids <- c("ENST00000234420", "ENST00000269305", "ENST00000445888")
#'
#' PrepareAnnotationEnsembl2(mart=ensembl, annotation_path=annotation_path,
#' splice_matrix=FALSE, dbsnp=NULL, transcript_ids=transcript_ids,
#' COSMIC=FALSE)
PrepareAnnotationEnsembl2 <- function(mart, annotation_path,
splice_matrix=FALSE, dbsnp=NULL,
transcript_ids=NULL, COSMIC=FALSE,...) {
options(stringsAsFactors=FALSE)
dataset <- biomaRt:::martDataset(mart)
biomart <- biomaRt:::martBM(mart)
host <- strsplit(strsplit(biomaRt:::martHost(mart),':')[[1]][2],'//')[[1]][2]
if (!is.null(dbsnp)) {
session <- browserSession() #
if(dataset == 'hsapiens_gene_ensembl') {
if(host == 'may2009.archive.ensembl.org'){
genome(session) <- 'hg18'
dbsnps <- 'snp130'
}else{
genome(session) <- 'hg19'
dbsnps <- trackNames(session)[grep('snp', trackNames(session),
fixed=TRUE)]
}
}
if(dataset == 'mmusculus_gene_ensembl') {
if(host == 'jan2013.archive.ensembl.org' ||
host == 'oct2012.archive.ensembl.org'){
genome(session) <- 'mm10'
dbsnps <- 'snp137'
}else{
genome(session) <- 'mm9'
dbsnps <- 'snp128'
}
}
dbsnp <- pmatch(dbsnp, dbsnps)#
if (is.na(dbsnp))
stop("invalid dbsnp name for specified genome")
if (dbsnp == -1)
stop("ambiguous dbsnp name")
}
message("Prepare gene/transcript/protein id mapping information (ids.RData) ... ",
appendLF=FALSE)
if(is.null(transcript_ids)){
transcript_ids <- getBM(attributes=c("ensembl_transcript_id"),
mart=mart)[,1]
}
attributes.id <- c("ensembl_gene_id","description")
idstab <- getBM(attributes=attributes.id, mart=mart,
filters='ensembl_transcript_id', values=transcript_ids)
colnames(idstab) <- c("ensembl_gene_id","description")
attributes.tr <- c("ensembl_gene_id", "ensembl_transcript_id",
"ensembl_peptide_id")
tr <- getBM(attributes=attributes.tr, mart=mart,
filters='ensembl_transcript_id', values=transcript_ids)
colnames(tr) <- c("ensembl_gene_id", "ensembl_transcript_id",
"ensembl_peptide_id")
ids <- merge(tr, idstab, by='ensembl_gene_id')
colnames(ids) <- c('gene_name', 'tx_name', 'pro_name', 'description')
save(ids, file=paste(annotation_path, '/ids.RData', sep=''))
packageStartupMessage(" done")
message("Build TranscriptDB object (txdb.sqlite) ... ", appendLF=TRUE)
tr_coding <- subset(ids, pro_name != "")
tr_noncoding <- subset(ids, pro_name == "")
txdb<- customProDB:::makeTranscriptDbFromBiomart_archive(biomart=biomart,
dataset=dataset,
host=host,
path="/biomart/martservice",
archive=FALSE,
transcript_ids=transcript_ids)
#saveFeatures(txdb, file=paste(annotation_path,'/txdb.sqlite',sep=''))
saveDb(txdb, file=paste(annotation_path, '/txdb.sqlite', sep=''))
packageStartupMessage(" done")
#txdb_coding <- makeTranscriptDbFromBiomart_archive(biomart=biomart,
# dataset=dataset, host=host, path="/biomart/martservice", archive=FALSE,
# transcript_ids=tr_coding[, "tx_name"])
#saveFeatures(txdb_coding, file=paste(annotation_path, '/txdb_coding.sqlite', sep=''))
#saveDb(txdb_coding, file=paste(annotation_path, '/txdb_coding.sqlite', sep=''))
#txdb_noncoding <- makeTranscriptDbFromBiomart_archive(biomart=biomart,
# dataset =dataset, host=host, path="/biomart/martservice",
# archive=FALSE, transcript_ids=tr_noncoding[, "tx_name"])
#saveFeatures(txdb_noncoding, file=paste(annotation_path, '/txdb_noncoding.sqlite', sep=''))
#saveDb(txdb_noncoding, file=paste(annotation_path, '/txdb_noncoding.sqlite', sep=''))
transGrange <- transcripts(txdb)
transintxdb <- IRanges::as.data.frame(transGrange)[, c('tx_id', 'tx_name')]
#exon_anno.RData
message("Prepare exon annotation information (exon_anno.RData) ... ",
appendLF=FALSE)
attributes.exon <- c("ensembl_exon_id", "ensembl_peptide_id",
'ensembl_gene_id', "chromosome_name",
"start_position", "end_position",
"exon_chrom_start", "exon_chrom_end", "strand",
"5_utr_start", "5_utr_end", "3_utr_start",
"3_utr_end", "cds_start", "cds_end", "rank",
"ensembl_transcript_id")
# download exon information
exon <- getBM(attributes=attributes.exon, mart=mart,
filters='ensembl_transcript_id', values=transcript_ids)
colnames(exon) <- attributes.exon
exon <- merge(exon, transintxdb, by.x="ensembl_transcript_id",
by.y="tx_name")
colnames(exon) <- c("tx_name", "exon_name", "pro_name", "gene_name",
"chromosome_name", "start_position", "end_position",
"exon_chrom_start", "exon_chrom_end", "strand",
"5_utr_start", "5_utr_end", "3_utr_start",
"3_utr_end", "cds_start", "cds_end", "rank", "tx_id")
# cdsBy is a function from package "GenomicFeatures".
# It's used to extract the cds from txdb.
cdsByTx <- cdsBy(txdb, "tx", use.names=TRUE)
# convert object of IRanges to data.frame
cdss <- IRanges::as.data.frame(cdsByTx)
cds_chr_p <- data.frame(tx_name=cdss[, "group_name"],
cds_chr_start=cdss[, "start"],
cds_chr_end=cdss[, "end"],
rank=cdss[, "exon_rank"])
# substitude "group" with "group_name", modified by Shawn,"group" was just
# supported by the version before bioconductor 3.1.
cds_chr_p_coding <- subset(cds_chr_p, tx_name %in% tr_coding[, 'tx_name'])
exon <- merge(exon, cds_chr_p_coding, by.y=c("tx_name", "rank"),
by.x=c("tx_name", "rank"), all.x=TRUE)
save(exon,file=paste(annotation_path, '/exon_anno.RData', sep=''))
packageStartupMessage(" done")
# procodingseq.RData is not useful for AS construction.
message("Prepare protein coding sequence (procodingseq.RData)... ",
appendLF=FALSE)
attributes.codingseq <- c("coding", "ensembl_peptide_id",
"ensembl_transcript_id")
if(length(tr_coding[, 'pro_name']<10000)){
coding <- getBM(attributes=attributes.codingseq,
filters="ensembl_peptide_id",
values=tr_coding[, 'pro_name'],
mart=mart)
}else{
index <- floor(length(tr_coding[, 'pro_name'])/10000)
coding <- c()
for(i in 1:index) {
st <- (i-1)*10000+1
ed <- i*10000
tmp <- getBM(attributes=attributes.codingseq,
filters="ensembl_peptide_id",
values=tr_coding[st:ed, 'pro_name'], mart=mart)
coding <- rbind(coding, tmp)
#print(i)
}
tmp <- getBM(attributes=attributes.codingseq,
filters="ensembl_peptide_id",
values=tr_coding[ed+1:length(tr_coding[, 'pro_name']),
'pro_name'],
mart=mart)
coding <- rbind(coding, tmp)
}
colnames(coding) <- attributes.codingseq
tx_id <- transintxdb[match(coding[, 'ensembl_transcript_id'],
transintxdb[, 'tx_name']),
'tx_id']
procodingseq <- cbind(coding, tx_id)
colnames(procodingseq) <- c("coding", "pro_name", "tx_name", "tx_id")
save(procodingseq,file=paste(annotation_path,'/procodingseq.RData',sep=''))
packageStartupMessage(" done")
message("Prepare protein sequence (proseq.RData) ... ", appendLF=FALSE)
attributes.proseq <- c("peptide", "ensembl_peptide_id",
"ensembl_transcript_id")
if(length(tr_coding[, 'pro_name']<10000)){
proteinseq <- getBM(attributes=attributes.proseq,
filters="ensembl_peptide_id",
values=tr_coding[,'pro_name'], mart=mart)
}else{
index <- floor(length(tr_coding[, 'pro_name'])/10000)
proteinseq <- c()
for(i in 1:index) {
st <- (i-1)*10000+1
ed <- i*10000
tmp <- getBM(attributes=attributes.proseq,
filters="ensembl_peptide_id",
values= tr_coding[st:ed, 'pro_name'],
mart=mart)
proteinseq <- rbind(proteinseq, tmp)
#print(i)
}
tmp <- getBM(attributes=attributes.proseq, filters="ensembl_peptide_id",
values=tr_coding[ed+1:length(tr_coding[, 'pro_name']),
'pro_name'],
mart=mart)
proteinseq <- rbind(proteinseq, tmp)
}
colnames(proteinseq) <- c("peptide", "pro_name", "tx_name")
save(proteinseq, file=paste(annotation_path, '/proseq.RData', sep=''))
packageStartupMessage(" done")
if (!is.null(dbsnp)) {
message("Prepare dbSNP information (dbsnpinCoding.RData) ... ",
appendLF=FALSE)
if(length(dbsnps) == 1&&dbsnps == 'snp128'){
dbsnp_query <- ucscTableQuery(session, dbsnps[dbsnp],
table='snp128')
}else{
dbsnp_query <- ucscTableQuery(session, dbsnps[dbsnp],
table=paste(dbsnps[dbsnp],
'CodingDbSnp', sep=''))
}
snpCodingTab <- getTable(dbsnp_query)
snpCodingTab[, 'chrom'] <- gsub('chr', '', snpCodingTab[, 'chrom'])
chrlist <- paste(c(seq(1:22),'X','Y'))
snpCoding <- subset(snpCodingTab, chrom %in% chrlist ,
select=c(chrom:name, alleleCount, alleles))
snpCoding <- unique(snpCoding)
#snpCoding[, 'chrom'] <- gsub('chrM', 'MT', snpCoding[, 'chrom'])
#
#save(snpCoding,file=paste(annotation_path,'/snpcoding.RData',sep=''))
snpCoding <- GRanges(seqnames=snpCoding[, 'chrom'],
ranges=IRanges(start=snpCoding[, 'chromStart'],
end=snpCoding[, 'chromEnd']),
strand='*',
rsid=snpCoding[, 'name'],
alleleCount=snpCoding[, 'alleleCount'],
alleles=snpCoding[, 'alleles'])
#seqlevels(snpCoding)
#if(TRUE%in% grep('chr',seqlevels(snpCoding)) > 0 ) {
# rchar <- sub('chr','',seqlevels(snpCoding))
# names(rchar) <- seqlevels(snpCoding)
# snpCoding <- renameSeqlevels(snpCoding, rchar) }
#if('M'%in%seqlevels(snpCoding)) snpCoding <- renameSeqlevels(snpCoding, c( M='MT'))
#chrlist <- paste(c(seq(1:22),'X','Y'))
transGrange_snp <- transGrange
#transGrange_snp <- keepSeqlevels(transGrange_snp, snpCoding)
#snpCoding <- keepSeqlevels(snpCoding, transGrange_snp)
#snpCoding <- keepSeqlevels(snpCoding, transGrange)
dbsnpinCoding <- subsetByOverlaps(snpCoding,transGrange_snp)
save(dbsnpinCoding,file=paste(annotation_path, '/dbsnpinCoding.RData',
sep=''))
packageStartupMessage(" done")
}
if (COSMIC) {
message("Prepare COSMIC information (cosmic.RData) ... ",
appendLF=FALSE)
cosmic_query <- ucscTableQuery(session, 'cosmic', table='cosmic')
cosmicTab <- getTable(cosmic_query)
cosmicTab[,'chrom'] <- gsub('chrM', 'MT', cosmicTab[, 'chrom'])
cosmicTab[,'chrom'] <- gsub('chr', '', cosmicTab[, 'chrom'])
chrlist <- paste(c(seq(1:22),'X','Y','MT'))
cosmicTab <- subset(cosmicTab, chrom %in% chrlist)
cosmic <- GRanges(seqnames=cosmicTab[, 'chrom'],
ranges=IRanges(start=cosmicTab[, 'chromStart'],
end=cosmicTab[, 'chromEnd']),
strand = '*',
cosid=cosmicTab[, 'name'])
transGrange_cosmic <- transGrange
#transGrange_cosmic <- keepSeqlevels(transGrange_cosmic, cosmic)
#cosmic <- keepSeqlevels(cosmic, transGrange_cosmic)
cosmic <- subsetByOverlaps(cosmic, transGrange_cosmic)
save(cosmic,file=paste(annotation_path, '/cosmic.RData', sep=''))
packageStartupMessage(" done")
}
if(splice_matrix){
message("Prepare exon splice information (splicemax.RData) ... ",
appendLF=FALSE)
exonByTx <- exonsBy(txdb, "tx", use.names=FALSE)
index <- which(elementNROWS(exonByTx)==1)
exonByTx_mul <- exonByTx[-index]
exons_mul <- IRanges::as.data.frame(exonByTx_mul)
exonslist <- split(exons_mul, exons_mul$group)
splicemax_list <- lapply(exonslist, .gen_splicmatrix)
splicemax <- do.call(rbind, splicemax_list)
save(splicemax, file=paste(annotation_path, '/splicemax.RData', sep=''))
packageStartupMessage(" done")
}
#splice <- paste(splicemax[, 1], splicemax[, 2], sep='-')
}
#' @title Prepare annotation from Refseq
#' @description Prepare the annotation for Refseq through UCSC table browser. This
#' function is modified from the function \code{\link{PrepareAnnotationRefseq}} in \pkg{customProDB}.
#' @param genome See detail in function \code{\link{PrepareAnnotationRefseq}}.
#' @param CDSfasta See detail in function \code{\link{PrepareAnnotationRefseq}}.
#' @param pepfasta See detail in function \code{\link{PrepareAnnotationRefseq}}.
#' @param annotation_path See detail in function \code{\link{PrepareAnnotationRefseq}}.
#' @param dbsnp See detail in function \code{\link{PrepareAnnotationRefseq}}.
#' @param transcript_ids See detail in function \code{\link{PrepareAnnotationRefseq}}.
#' @param splice_matrix See detail in function \code{\link{PrepareAnnotationRefseq}}.
#' @param COSMIC See detail in function \code{\link{PrepareAnnotationRefseq}}.
#' @param ... Additional arguments
#' @return Several .RData file containing annotations needed for following analysis.
#' @seealso \code{\link{PrepareAnnotationEnsembl2}}.
#' @export
#' @examples
#' \dontrun{
#' transcript_ids <- c("NM_001126112", "NM_033360", "NR_073499")
#' pepfasta <- system.file("extdata", "refseq_pro_seq.fasta",
#' package="customProDB")
#' CDSfasta <- system.file("extdata", "refseq_coding_seq.fasta",
#' package="customProDB")
#' annotation_path <- tempdir()
#' PrepareAnnotationRefseq2(genome='hg19', CDSfasta, pepfasta, annotation_path,
#' dbsnp=NULL, transcript_ids=transcript_ids,
#' splice_matrix=FALSE, COSMIC=FALSE)
#' }
PrepareAnnotationRefseq2 <- function(genome='hg19', CDSfasta, pepfasta,
annotation_path, dbsnp=NULL,
transcript_ids=NULL, splice_matrix=FALSE,
COSMIC=FALSE, ...) {
options(stringsAsFactors=FALSE)
session <- browserSession()
genome(session) <- genome
tablename <- 'refGene'
message("Build TranscriptDB object (txdb.sqlite) ... ", appendLF=TRUE)
txdb <- makeTxDbFromUCSC(genome=genome, tablename=tablename,
transcript_ids=transcript_ids)
saveDb(txdb, file=paste(annotation_path, '/txdb.sqlite', sep=''))
packageStartupMessage(" done")
message("Prepare gene/transcript/protein id mapping information (ids.RData) ... ",
appendLF=FALSE)
query_refGene <- ucscTableQuery(session, "refSeqComposite", table="refGene")
refGene <- getTable(query_refGene)
if(!is.null(transcript_ids))
{
refGene <- subset(refGene, name %in% transcript_ids)
}
reflink <- .UCSC_dbselect("hgFixed", "refLink")
ids <- subset(reflink, mrnaAcc %in% refGene[, 'name'], select = name:protAcc)
colnames(ids) <- c('gene_name', 'description', 'tx_name', 'pro_name')
save(ids, file=paste(annotation_path, '/ids.RData', sep=''))
packageStartupMessage(" done")
#tr_coding <- subset(ids,pro_name!="")
#tr_noncoding <- subset(ids,pro_name == "")
#txdb_coding <- makeTranscriptDbFromUCSC(genome=genome, tablename=tablename,
# transcript_ids=tr_coding[, "tx_name"] )
#saveDb(txdb_coding,
# file=paste(annotation_path, '/txdb_coding.sqlite', sep=''))
#txdb_noncoding <- makeTranscriptDbFromUCSC(genome=genome,
# tablename=tablename, transcript_ids=tr_noncoding[, "tx_name"] )
#saveDb(txdb_noncoding,
# file=paste(annotation_path, '/txdb_noncoding.sqlite', sep=''))
message("Prepare exon annotation information (exon_anno.RData) ... ",
appendLF=FALSE)
transGrange <- transcripts(txdb)
tr <- IRanges::as.data.frame(transGrange)
cdsByTx <- cdsBy(txdb, "tx", use.names=FALSE)
exonByTx <- exonsBy(txdb, "tx", use.names=FALSE)
fiveutrByTx <- fiveUTRsByTranscript(txdb, use.names=FALSE)
threeutrByTx <- threeUTRsByTranscript(txdb, use.names=FALSE)
#####################################################
# get error in most recent version of IRanges package
# previous: rownames after unlist 1.1 1.2 1.3
# now: .1 .2 .3 ... were removed, so there are some rows with same rownames
######################################################
#cdss <- IRanges::as.data.frame(IRanges::unlist(cdsByTx))
#exons <- IRanges::as.data.frame(IRanges::unlist(exonByTx))
#fiveutrs <- IRanges::as.data.frame(IRanges::unlist(fiveutrByTx))
#threeutrs <- IRanges::as.data.frame(IRanges::unlist(threeutrByTx))
cdss <- IRanges::as.data.frame(cdsByTx)
exons <- IRanges::as.data.frame(exonByTx)
fiveutrs <- IRanges::as.data.frame(fiveutrByTx)
threeutrs <- IRanges::as.data.frame(threeutrByTx)
#txid <- matrix(unlist(strsplit(rownames(exons), '\\.')), ncol = 2,
#byrow =T)[, 1]
#txid <- gsub('=','\\.', txid)
#substitude "group" with "group_name", modified by Shawn."group" was just
#supported by the version before bioconductor 3.1
exon_p <- data.frame(txid=exons[, "group_name"], chr=exons[, "seqnames"],
exon_s=exons[, "start"], exon_e=exons[, "end"],
exon_rank=exons[, "exon_rank"])
exon2tr <- merge(exon_p, tr,by.y="tx_id", by.x="txid")
exon2tr <- exon2tr[, -which(names(exon2tr) %in% c("seqnames", "width"))]
#txid <- matrix(unlist(strsplit(rownames(cdss), '\\.')), ncol = 2,
# byrow =T)[, 1]
#txid <- gsub('=','\\.',txid)
#substitude "group" with "group_name", modified by Shawn
cds_p <- data.frame(txid=cdss[, "group_name"], cds_s=cdss[, "start"],
cds_e=cdss[, "end"], exon_rank=cdss[, "exon_rank"],
width=cdss[, "width"])
ttt <- split(cds_p, cds_p$txid)
cds_p_new_list <-lapply(ttt, function(x){
#len <- x[,'cds_e']-x[,'cds_s']+1
#cum <- cumsum(len)
cum <- cumsum(x[, 'width'])
rdis <- cbind(c(1, cum[1:length(cum)-1]+1), cum)
colnames(rdis) <- c('cds_start', 'cds_end')
tmp <- cbind(x, rdis)
tmp
})
cds_p_new <- do.call(rbind, cds_p_new_list)
cds_p_new <- cds_p_new[, -which(names(cds_p_new) %in% c("width"))]
#for(i in 1:length(ttt)) {
# print(i)
# ttt1 <- ttt[[i]]
# len <- ttt1[,'cds_e']-ttt1[,'cds_s']+1
# cum <- cumsum(len)
# rdis <- cbind(c(1,cum[1:length(cum)-1]+1),cum)
# colnames(rdis) <- c('cds_start','cds_end')
# tmp <- cbind(ttt1,rdis)
# cds_p_new <- rbind(cds_p_new,tmp)
#}
cds2exon <- merge(exon2tr, cds_p_new, by.x=c("txid", "exon_rank"),
by.y=c("txid", "exon_rank"), all.x = TRUE)
#txid <- matrix(unlist(strsplit(rownames(fiveutrs), '\\.')), ncol = 2,
# byrow =T)[, 1]
#txid <- gsub('=','\\.', txid)
#substitude "group" with "group_name", modified by Shawn
fiveutr_p <- data.frame(txid=fiveutrs[, "group_name"],
fiveutr_s=fiveutrs[, "start"],
fiveutr_e=fiveutrs[, "end"],
exon_rank=fiveutrs[, "exon_rank"])
fiveutr2exon <- merge(cds2exon, fiveutr_p, by.x=c("txid", "exon_rank"),
by.y =c("txid", "exon_rank"), all.x = TRUE)
#txid <- matrix(unlist(strsplit(rownames(threeutrs),'\\.')),
#ncol = 2,byrow =T)[, 1]
#txid <- gsub('=','\\.', txid)
#substitude "group" with "group_name", modified by Shawn
threeutr_p <- data.frame(txid=threeutrs[, "group_name"],
threeutr_s=threeutrs[, "start"],
threeutr_e=threeutrs[, "end"],
exon_rank=threeutrs[, "exon_rank"])
threeutr2exon <- merge(fiveutr2exon, threeutr_p,
by.x=c("txid", "exon_rank"),
by.y=c("txid", "exon_rank"), all.x = TRUE)
#exon <- merge(threeutr2exon,ids,by.x=c("tx_name"),by.y="tx_name",
#all.x = TRUE)
exon <- threeutr2exon[order(threeutr2exon$txid, threeutr2exon$exon_rank), ]
colnames(exon) <- c("tx_id","rank", "chromosome_name", "exon_chrom_start",
"exon_chrom_end", "start_position", "end_position",
"strand", "tx_name", "cds_chr_start", "cds_chr_end",
"cds_start", "cds_end", "5_utr_start",
"5_utr_end", "3_utr_start", "3_utr_end")
pro_name <- ids[match(exon[, 'tx_name'], ids[, 'tx_name']), 'pro_name']
gene_name <- ids[match(exon[, 'tx_name'], ids[, 'tx_name']), 'gene_name']
exon <- cbind(exon, pro_name, gene_name)
save(exon, file=paste(annotation_path, '/exon_anno.RData', sep=''))
packageStartupMessage(" done")
message("Prepare protein sequence (proseq.RData) ... ", appendLF=FALSE)
pro_seqs <- readAAStringSet(pepfasta, format= 'fasta')
pro_name_v <- names(pro_seqs)
pro_name <- unlist(lapply(pro_name_v, function(x) strsplit(x, '\\.')[[1]][1]))
tx_name <- ids[match(pro_name, ids[, 'pro_name']), 'tx_name']
proteinseq <- as.data.frame(pro_seqs)
proteinseq <- cbind(proteinseq, pro_name_v, pro_name, tx_name)
colnames(proteinseq) <- c("peptide", "pro_name_v", "pro_name", "tx_name")
proteinseq <- subset(proteinseq, tx_name %in% refGene[, 'name'])
save(proteinseq, file=paste(annotation_path, '/proseq.RData', sep=''))
packageStartupMessage(" done")
message("Prepare protein coding sequence (procodingseq.RData)... ",
appendLF=FALSE)
cds_seqs <- readDNAStringSet(CDSfasta, format= 'fasta')
tx_name_tmp <- unlist(lapply(names(cds_seqs), function(x)
strsplit(x, ' ')[[1]][1]))
tx_name <- unlist(lapply(tx_name_tmp, function(x)
paste(strsplit(x, '_')[[1]][3:4], collapse='_')))
tx_range_tmp <- unlist(lapply(names(cds_seqs), function(x)
strsplit(x, ' ')[[1]][2]))
tx_chr <- unlist(lapply(tx_range_tmp, function(x)
substring(strsplit(x, ':')[[1]][1],7)))
tx_cds_sta <- unlist(lapply(tx_range_tmp, function(x)
strsplit(strsplit(x, ':')[[1]][2], '-')[[1]][[1]]))
tx_cds_end <- unlist(lapply(tx_range_tmp, function(x)
strsplit(strsplit(x, ':')[[1]][2], '-')[[1]][[2]]))
tx_name_cds <- refGene[match(paste(tx_name, tx_chr, tx_cds_sta, tx_cds_end,
sep=' '),
paste(refGene[, 'name'], refGene[, 'chrom'],
refGene[, 'cdsStart']+1,
refGene[, 'cdsEnd'], sep=' ')),
c('name','chrom','txStart','txEnd')]
tx_id <- tr[match(paste(tx_name_cds[, 'name'], tx_name_cds[, 'chrom'],
tx_name_cds[, 'txStart']+1,
tx_name_cds[, 'txEnd'],
sep=' '),
paste(tr[, 'tx_name'], tr[, 'seqnames'], tr[, 'start'],
tr[, 'end'], sep=' ')), 'tx_id']
pro_name <- ids[match(tx_name,ids[, 'tx_name']), 'pro_name']
procodingseq <- as.data.frame(cds_seqs)
procodingseq <- cbind(procodingseq, names(cds_seqs), pro_name,
tx_name, tx_id)
colnames(procodingseq) <- c("coding", "tx_name_full", "pro_name",
"tx_name", "tx_id")
procodingseq <- subset(procodingseq, tx_name %in% refGene[, 'name'])
save(procodingseq, file=paste(annotation_path, '/procodingseq.RData',
sep=''))
packageStartupMessage(" done")
if (!is.null(dbsnp)) {
message("Prepare dbSNP information (dbsnpinCoding.RData) ... ",
appendLF=FALSE)
dbsnps <- trackNames(session)[grep('snp', trackNames(session),
fixed=TRUE)]
dbsnp <- pmatch(dbsnp, dbsnps)
if (is.na(dbsnp))
stop("invalid dbsnp name for specified genome")
if (dbsnp == -1)
stop("ambiguous dbsnp name")
dbsnp_query <- ucscTableQuery(session, dbsnps[dbsnp],
table=paste(dbsnps[dbsnp],
'CodingDbSnp', sep=''))
snpCodingTab <- getTable(dbsnp_query)
snpCoding <- subset(snpCodingTab,transcript %in% refGene[, 'name'],
select=c(chrom:name, alleleCount, alleles))
snpCoding <- unique(snpCoding)
#save(snpCoding,file=paste(annotation_path, '/snpcoding.RData', sep=''))
dbsnpinCoding <- GRanges(seqnames=snpCoding[, 'chrom'],
ranges=IRanges(start=snpCoding[, 'chromStart'],
end=snpCoding[, 'chromEnd']),
strand='*',
rsid=snpCoding[, 'name'],
alleleCount=snpCoding[, 'alleleCount'],
alleles=snpCoding[, 'alleles'])
save(dbsnpinCoding, file=paste(annotation_path, '/dbsnpinCoding.RData',
sep=''))
packageStartupMessage(" done")
}
if (COSMIC) {
#cosmic <- trackNames(session)[grep('cosmic',trackNames(session),
#fixed=T)]
message("Prepare COSMIC information (cosmic.RData) ...",appendLF=FALSE)
cosmic_query <- ucscTableQuery(session, 'cosmic', table='cosmic')
cosmicTab <- getTable(cosmic_query)
cosmic <- GRanges(seqnames=cosmicTab[, 'chrom'],
ranges=IRanges(start=cosmicTab[, 'chromStart'],
end=cosmicTab[, 'chromEnd']),
strand = '*', cosid=cosmicTab[,'name'])
#cosmic <- keepSeqlevels(cosmic,transGrange)
cosmic <- subsetByOverlaps(cosmic, transGrange)
save(cosmic,file=paste(annotation_path, '/cosmic.RData', sep=''))
packageStartupMessage(" done")
}
if(splice_matrix){
message("Prepare exon splice information (splicemax.RData) ... ",
appendLF=FALSE)
index <- which(elementNROWS(exonByTx)==1)
exonByTx_mul <- exonByTx[-index]
exons_mul <- IRanges::as.data.frame(exonByTx_mul)
exonslist <- split(exons_mul, exons_mul$group)
#system.time( exonByTx <- exonsBy(txdb,"tx", use.names=F))
splicemax_list <- lapply(exonslist, .gen_splicmatrix)
splicemax <- do.call(rbind, splicemax_list)
save(splicemax, file=paste(annotation_path, '/splicemax.RData',
sep=''))
packageStartupMessage(" done")
}
}
.gen_splicmatrix <- function(x, ...) {
mystrand=x[1,'strand']
a=x[,'exon_rank']
b=x[,'exon_id']
n=length(a)
if (mystrand=='+'){
tmp=order(a)
}else{
tmp=order(a,decreasing=TRUE)
}
mm=cbind(b[tmp[1:(n-1)]], b[tmp[2:n]])
return(mm)
}
##' @title Create protein database for novel transcripts
##' @description Create protein database for novel transcripts
##' @param gffFile A GFF format file containing novel transcripts information
##' @param outmtab A txt format file containing the novel transcripts information
##' @param outfa The output fasta format protein sequence file
##' @param outgtf The output GTF format file
##' @param bool_get_longest When it's set as TRUE, the longest sequences will be
##' retained after the DNA sequences are six-frame translated into protein
##' sequences. Otherwise, the protein sequences more than 30 aa are retained.
##' @param organism What is the Genus and species of this organism.Please use
##' proper scientific nomenclature for example: "Homo sapiens" and not "human",
##' default is "Homo sapiens".
##' @param genome Genome information
##' @param ... Additional arguments
##' @return The database file(s)
##' @noRd
getNovelTx=function(gffFile="transcripts.gtf",
outmtab="novel_transcripts_ntx.tab",
outfa="novel_transcripts_ntx.fasta",
outgtf="novel_transcripts_ntx.gtf",
bool_get_longest=TRUE,genome=NULL,organism="Homo sapiens"){
options(stringsAsFactors = FALSE)
if(is.null(genome)){
stop("genome not found!")
}
########## Params
if(file.exists(outfa)){
file.remove(outfa)
}
tmpdir<-tempdir()
chromLen<-seqlengths(genome)
#######filter
gff <- import(gffFile)
novel_gff<-gff[substr(gff$transcript_id,1,5) =="CUFF."]
#justify the chromosomes are ucsc format or ensembl format
if(any(c(1:22, "X", "Y") %in% seqlevels(novel_gff))){
# restricting this extraction to the 24 chromosomes
seqlevels(novel_gff, pruning.mode="coarse") <- c(1:22, "X", "Y")
seqlevels(novel_gff) <- paste0("chr", seqlevels(novel_gff)) # rename
}else{
seqlevels(novel_gff, pruning.mode="coarse") <-
paste("chr",c(1:22, "X", "Y"), sep="")
}
novel_tx<-novel_gff[novel_gff$type=="transcript"]
#record the strand info
#dict_strand<-dlply(as.data.frame(novel_tx),.(transcript_id),
# summarize,strand=strand)
vector_nonstrand <- novel_tx[strand(novel_tx)=="*"]$transcript_id
vector_forward <- novel_tx[strand(novel_tx)=="+"]$transcript_id
vector_reverse <- novel_tx[strand(novel_tx)=="-"]$transcript_id
#the coordinate info can be found here.
export(novel_gff, outgtf, format="gtf")
strand(novel_gff)<-"+" #change the strand
tmpgff<-paste(tmpdir,"novel_transcripts_strand_forward.gtf",sep="/")
export(novel_gff, tmpgff, format="gtf")
#cat(paste("temp dir:",tmpdir,""sep=""))
#chrominfo<-data.frame(names(chromLen),as.vector(chromLen),
# rep(FALSE,length(chromLen)))
#colnames(chrominfo)<-c("chrom","length","is_circular")
# makeTxDbFromGFF {GenomicFeatures}
# Make a TxDb object from annotations available as a GFF3 or GTF file
#txdb <- makeTxDbFromGFF(file=tmpgff,format="gtf",
# exonRankAttributeName="exon_number",
# species=species,
# circ_seqs=names(chromLen))
txdb <- makeTxDbFromGFF(file=tmpgff,format="gtf",
#exonRankAttributeName="exon_number",
organism=organism)
#if(interactive()) {
#saveDb(txdb,file="TESTGTF.sqlite")
#}
## Extract the exon ranges grouped by transcript from 'txdb':
transcripts <- exonsBy(txdb, by="tx", use.names=TRUE)
#justify the chromosomes are ucsc format or ensembl format
#if(any(c(1:22, "X", "Y") %in% seqlevels(transcripts)))
#{
# restricting this extraction to the 24 chromosomes
#seqlevels(transcripts, pruning.mode="coarse") <- c(1:22, "X", "Y")
#seqlevels(transcripts) <- paste0("chr", seqlevels(transcripts)) # rename
#}
## Extract the transcript sequences from the genome:
tx_seqs <- extractTranscriptSeqs(genome, transcripts)
cumIndex <-0
if(length(vector_reverse)>0)
{
message("====== Processing reverse strand ======")
tx_seqs_reverse<-tx_seqs[match(vector_reverse,names(tx_seqs))]
tx_seqs_normal<-reverseComplement(tx_seqs_reverse)
peptides_r1<-suppressWarnings(translate(tx_seqs_normal,
if.fuzzy.codon="solve"))
peptides_r2<-suppressWarnings(translate(subseq(tx_seqs_normal,start=2),
if.fuzzy.codon="solve"))
peptides_r3<-suppressWarnings(translate(subseq(tx_seqs_normal,start=3),
if.fuzzy.codon="solve"))
#tmp_vector<-unlist(strsplit(as.data.frame(peptides_r1)$x, "\\*"))
#way1: find the longest sequence
#tmp_vector[str_length(tmp_vector)==max(str_length(tmp_vector))]
#way2: find all of the sequences more than 30 AA
#tmp_vector[str_length(tmp_vector)>=30]
novel_cds_r1 <- .get_30aa_splited_seq(peptides_r1)
novel_cds_r2 <- .get_30aa_splited_seq(peptides_r2)
novel_cds_r3 <- .get_30aa_splited_seq(peptides_r3)
novel_cds_r1[,c("Strand","Frame","ID"):=list(a="-",
b="1",
c=paste(id,"-","F1",
Substring,
sep="|")),
by=.(id,Substring)]
novel_cds_r2[,c("Strand","Frame","ID"):=list(a="-",
b="2",
c=paste(id,"-","F2",
Substring,
sep="|")),
by=.(id,Substring)]
novel_cds_r3[,c("Strand","Frame","ID"):=list(a="-",
b="3",
c=paste(id,"-","F3",
Substring,
sep="|")),
by=.(id,Substring)]
all_pep<-rbindlist(list(novel_cds_r1,novel_cds_r2,novel_cds_r3))
all_pep[,Index:=paste("NTX",cumIndex+.I,sep="")]
cumIndex<-dim(all_pep)[1]+cumIndex;
#all_pep<-rbind(cbind(names_r1,novel_cds_r1[,"subpep.seq"]),
#cbind(names_r2,novel_cds_r2[,"subpep.seq"]),
#cbind(names_r3,novel_cds_r3[,"subpep.seq"]))
if(bool_get_longest){
final_all_pep<-all_pep[,.(pep=seq[nchar(seq)==max(nchar(seq))],
ID=ID[nchar(seq)==max(nchar(seq))],
Index=Index[nchar(seq)==max(nchar(seq))],
Strand=Strand[nchar(seq)==max(nchar(seq))],
Frame=Frame[nchar(seq)==max(nchar(seq))],
start=start[nchar(seq)==max(nchar(seq))],
end=end[nchar(seq)==max(nchar(seq))],
Substring=Substring[nchar(seq)==max(nchar(seq))]),
by=.(id)]
#final_all_pep<-all_pep[,.(pep=seq[which.max(nchar(seq))],
#ID=ID[which.max(nchar(seq))]),by=.(id)]
write.table(subset(final_all_pep,
select=c(Index,id,Strand,Frame,start,end,Substring)),
file=outmtab, sep='\t', quote=FALSE, row.names=FALSE)
final_all_pep[,output:=paste('>',Index,"|",ID,'\n',pep,sep=''),
by=.(ID)]
write(final_all_pep$output,file=outfa,append=TRUE)
}else{
write.table(subset(all_pep,
select=c(Index,id,Strand,Frame,start,end,Substring)),
file=outmtab, sep='\t', quote=FALSE, row.names=FALSE)
all_pep[,output:=paste('>',Index,"|",ID,'\n',seq,sep=''),by=.(ID)]
write(all_pep$output,file=outfa,append=TRUE)
}
}
if(length(vector_forward)>0)
{
message("====== Processing forward strand ======")
tx_seqs_forward<-tx_seqs[match(vector_forward,names(tx_seqs))]
tx_seqs_normal<-tx_seqs_forward
peptides_f1<-suppressWarnings(translate(tx_seqs_normal,
if.fuzzy.codon="solve"))
peptides_f2<-suppressWarnings(translate(subseq(tx_seqs_normal,start=2),
if.fuzzy.codon="solve"))
peptides_f3<-suppressWarnings(translate(subseq(tx_seqs_normal,start=3),
if.fuzzy.codon="solve"))
#tmp_vector<-unlist(strsplit(as.data.frame(peptides_f1)$x, "\\*"))
#way1: find the longest sequence
#tmp_vector[str_length(tmp_vector)==max(str_length(tmp_vector))]
#way2: find all of the sequences more than 30 AA
#tmp_vector[str_length(tmp_vector)>=30]
novel_cds_f1 <- .get_30aa_splited_seq(peptides_f1)
novel_cds_f2 <- .get_30aa_splited_seq(peptides_f2)
novel_cds_f3 <- .get_30aa_splited_seq(peptides_f3)
novel_cds_f1[,c("Strand","Frame","ID"):=list(a="+",
b="1",
c=paste(id,"+","F1",
Substring,
sep="|")),
by=.(id,Substring)]
novel_cds_f2[,c("Strand","Frame","ID"):=list(a="+",
b="2",
c=paste(id,"+","F2",
Substring,
sep="|")),
by=.(id,Substring)]
novel_cds_f3[,c("Strand","Frame","ID"):=list(a="+",
b="3",
c=paste(id,"+","F3",
Substring,
sep="|")),
by=.(id,Substring)]
all_pep<-rbindlist(list(novel_cds_f1,novel_cds_f2,novel_cds_f3))
#all_pep<-rbind(cbind(names_f1,novel_cds_f1[,"subpep.seq"]),
#cbind(names_f2,novel_cds_f2[,"subpep.seq"]),
#cbind(names_f3,novel_cds_f3[,"subpep.seq"]))
all_pep[,Index:=paste("NTX",cumIndex+.I,sep="")]
cumIndex<-dim(all_pep)[1]+cumIndex;
if(bool_get_longest){
#final_all_pep<-all_pep[,.(pep=seq[which.max(nchar(seq))],
#ID=ID[which.max(nchar(seq))]),by=.(id)]
final_all_pep<-all_pep[,.(pep=seq[nchar(seq)==max(nchar(seq))],
ID=ID[nchar(seq)==max(nchar(seq))],
Index=Index[nchar(seq)==max(nchar(seq))],
Strand=Strand[nchar(seq)==max(nchar(seq))],
Frame=Frame[nchar(seq)==max(nchar(seq))],
start=start[nchar(seq)==max(nchar(seq))],
end=end[nchar(seq)==max(nchar(seq))],
Substring=Substring[nchar(seq)==max(nchar(seq))]),
by=.(id)]
if(file.exists(outfa)){
write.table(subset(final_all_pep,
select=c(Index,id,Strand,Frame,start,end,Substring)),
file=outmtab, sep='\t', quote=FALSE, row.names=FALSE,
append=TRUE,col.names=FALSE)
}else{
write.table(subset(final_all_pep,
select=c(Index,id,Strand,Frame,start,end,Substring)),
file=outmtab, sep='\t', quote=FALSE, row.names=FALSE)
}
final_all_pep[,output:=paste('>',Index,"|",ID,'\n',pep,sep=''),
by=.(ID)]
write(final_all_pep$output,file=outfa,append=TRUE)
}else{
if(file.exists(outfa)){
write.table(subset(all_pep,
select=c(Index,id,Strand,Frame,start,end,Substring)),
file=outmtab, sep='\t', quote=FALSE, row.names=FALSE,
append=TRUE,col.names=FALSE)
}else{
write.table(subset(all_pep,
select=c(Index,id,Strand,Frame,start,end,Substring)),
file=outmtab, sep='\t', quote=FALSE, row.names=FALSE)
}
all_pep[,output:=paste('>',Index,"|",ID,'\n',seq,sep=''),by=.(ID)]
write(all_pep$output,file=outfa,append=TRUE)
}
}
if(length(vector_nonstrand)>0){
message("====== Processing unknown strand ======")
tx_seqs_non<-tx_seqs[match(vector_nonstrand,names(tx_seqs))]
tx_seqs_normal<-tx_seqs_non
peptides_f1<-suppressWarnings(translate(tx_seqs_normal,
if.fuzzy.codon="solve"))
peptides_f2<-suppressWarnings(translate(subseq(tx_seqs_normal,start=2),
if.fuzzy.codon="solve"))
peptides_f3<-suppressWarnings(translate(subseq(tx_seqs_normal,start=3),
if.fuzzy.codon="solve"))
#tmp_vector<-unlist(strsplit(as.data.frame(peptides_f1)$x, "\\*"))
#way1: find the longest sequence
#tmp_vector[str_length(tmp_vector)==max(str_length(tmp_vector))]
#way2: find all of the sequences more than 30 AA
#tmp_vector[str_length(tmp_vector)>=30]
novel_cds_f1 <- .get_30aa_splited_seq(peptides_f1)
novel_cds_f2 <- .get_30aa_splited_seq(peptides_f2)
novel_cds_f3 <- .get_30aa_splited_seq(peptides_f3)
novel_cds_f1[,c("Strand","Frame","ID"):=list(a="+",
b="1",
c=paste(id,"+","F1",
Substring,
sep="|")),
by=.(id,Substring)]
novel_cds_f2[,c("Strand","Frame","ID"):=list(a="+",
b="2",
c=paste(id,"+","F2",
Substring,
sep="|")),
by=.(id,Substring)]
novel_cds_f3[,c("Strand","Frame","ID"):=list(a="+",
b="3",
c=paste(id,"+","F3",
Substring,
sep="|")),
by=.(id,Substring)]
############ reverseComplement
tx_seqs_normal<-reverseComplement(tx_seqs_non)
peptides_r1<-suppressWarnings(translate(tx_seqs_normal,
if.fuzzy.codon="solve"))
peptides_r2<-suppressWarnings(translate(subseq(tx_seqs_normal,start=2),
if.fuzzy.codon="solve"))
peptides_r3<-suppressWarnings(translate(subseq(tx_seqs_normal,start=3),
if.fuzzy.codon="solve"))
#tmp_vector<-unlist(strsplit(as.data.frame(peptides_r1)$x, "\\*"))
#way1: find the longest sequence
#tmp_vector[str_length(tmp_vector)==max(str_length(tmp_vector))]
#way2: find all of the sequences more than 30 AA
#tmp_vector[str_length(tmp_vector)>=30]
novel_cds_r1 <- .get_30aa_splited_seq(peptides_r1)
novel_cds_r2 <- .get_30aa_splited_seq(peptides_r2)
novel_cds_r3 <- .get_30aa_splited_seq(peptides_r3)
novel_cds_r1[,c("Strand","Frame","ID"):=list(a="-",
b="1",
c=paste(id,"-","F1",
Substring,
sep="|")),
by=.(id,Substring)]
novel_cds_r2[,c("Strand","Frame","ID"):=list(a="-",
b="2",
c=paste(id,"-","F2",
Substring,
sep="|")),
by=.(id,Substring)]
novel_cds_r3[,c("Strand","Frame","ID"):=list(a="-",
b="3",
c=paste(id,"-","F3",
Substring,
sep="|")),
by=.(id,Substring)]
all_pep<-rbindlist(list(novel_cds_f1,novel_cds_f2,novel_cds_f3,
novel_cds_r1,novel_cds_r2,novel_cds_r3))
all_pep[,Index:=paste("NTX",cumIndex+.I,sep="")]
if(bool_get_longest){
#final_all_pep<-all_pep[,.(pep=seq[nchar(seq)==max(nchar(seq))],
#ID=ID[nchar(seq)==max(nchar(seq))]),by=.(id)]
final_all_pep<-all_pep[,.(pep=seq[nchar(seq)==max(nchar(seq))],
ID=ID[nchar(seq)==max(nchar(seq))],
Index=Index[nchar(seq)==max(nchar(seq))],
Strand=Strand[nchar(seq)==max(nchar(seq))],
Frame=Frame[nchar(seq)==max(nchar(seq))],
start=start[nchar(seq)==max(nchar(seq))],
end=end[nchar(seq)==max(nchar(seq))],
Substring=Substring[nchar(seq)==max(nchar(seq))]),
by=.(id)]
if(file.exists(outmtab)){
write.table(subset(final_all_pep,
select=c(Index,id,Strand,Frame,start,end,Substring)),
file=outmtab, sep='\t', quote=FALSE, row.names=FALSE,
append=TRUE,col.names=FALSE)
}else{
write.table(subset(final_all_pep,
select=c(Index,id,Strand,Frame,start,end,Substring)),
file=outmtab, sep='\t', quote=FALSE, row.names=FALSE)
}
#final_all_pep<-all_pep[,.(pep=seq[which.max(nchar(seq))],ID=ID[which.max(nchar(seq))]),by=.(id)]
final_all_pep[,output:=paste('>',Index,"|",ID,'\n',pep,sep=''),
by=.(ID)]
write(final_all_pep$output,file=outfa,append=TRUE)
}else{
if(file.exists(outmtab)){
write.table(subset(all_pep,
select=c(Index,id,Strand,Frame,start,end,Substring)),
file=outmtab, sep='\t', quote=FALSE, row.names=FALSE,
append=TRUE,col.names=FALSE)
}else{
write.table(subset(all_pep,
select=c(Index,id,Strand,Frame,start,end,Substring)),
file=outmtab, sep='\t', quote=FALSE, row.names=FALSE)
}
all_pep[,output:=paste('>',Index,"|",ID,'\n',seq,sep=''),by=.(ID)]
write(all_pep$output,file=outfa,append=TRUE)
}
}
}
#' @title Create protein database based on the transcripts from de novo
#' reconstruction of transcriptome from RNA-seq data
#' @description Create protein database based on the transcripts from de novo
#' reconstruction of transcriptome from RNA-seq data.
#' @param infa A FASTA format file containing transcript sequences
#' (such as derived from de novo transcriptome assembly by Trinity)
#' @param bool_use_3frame A logical variable indicating whether to translate the
#' raw sequences with 3-frame (forward). Default is 6-frame translation (FALSE).
#' @param outmtab A txt format file containing the novel transcripts information
#' @param outfa The output fasta format protein sequence file
#' @param bool_get_longest When it's set as TRUE, only the longest protein sequences will be
#' retained after the raw sequences are translated with 3-frame or 6-frame.
#' Otherwise, all the protein sequences with longer than 30 aa will be retained.
#' @param make_decoy A logical variable indicating whether to add the decoy sequences.
#' @param decoy_tag The prefix of decoy sequence IDs.
#' @param outfile_name Output file name
#' @return The database file(s)
#' @export
#' @examples
#' transcript_seq_file <- system.file("extdata/input", "Trinity.fasta",package="PGA")
#' createProDB4DenovoRNASeq(infa=transcript_seq_file)
createProDB4DenovoRNASeq<-function(infa="./trinity.fasta",
bool_use_3frame=FALSE,
outmtab="novel_transcripts_ntx.tab",
outfa="./novel_transcripts_ntx.fasta",
bool_get_longest=TRUE,
make_decoy=TRUE,
decoy_tag="#REV#",
outfile_name="test"){
options(stringsAsFactors=FALSE)
if(file.exists(outfa)){
file.remove(outfa)
}
var_tag <- "DNO"
cumIndex <-0
dna_seq_forward<-readDNAStringSet(infa)
names(dna_seq_forward)<-gsub(" .+","",names(dna_seq_forward)) #get the id, remove the description
message("....... Proteomic database construction ! ............")
# forward strand
peptides_f1<-suppressWarnings(translate(dna_seq_forward,
if.fuzzy.codon="solve"))
peptides_f2<-suppressWarnings(translate(subseq(dna_seq_forward,start=2),
if.fuzzy.codon="solve"))
peptides_f3<-suppressWarnings(translate(subseq(dna_seq_forward,start=3),
if.fuzzy.codon="solve"))
novel_cds_f1 <- .get_30aa_splited_seq(peptides_f1)
novel_cds_f2 <- .get_30aa_splited_seq(peptides_f2)
novel_cds_f3 <- .get_30aa_splited_seq(peptides_f3)
novel_cds_f1[,c("Strand","Frame","ID"):=list(a="+",
b="1",
c=paste(id,"+","F1",
Substring,
sep="|")),
by=.(id,Substring)]
novel_cds_f2[,c("Strand","Frame","ID"):=list(a="+",
b="2",
c=paste(id,"+","F2",
Substring,
sep="|")),
by=.(id,Substring)]
novel_cds_f3[,c("Strand","Frame","ID"):=list(a="+",
b="3",
c=paste(id,"+","F3",
Substring,
sep="|")),
by=.(id,Substring)]
# reverse strand
if(!bool_use_3frame)
{
dna_seq_reverse<-reverseComplement(dna_seq_forward)
peptides_r1<-suppressWarnings(translate(dna_seq_reverse,
if.fuzzy.codon="solve"))
peptides_r2<-suppressWarnings(translate(subseq(dna_seq_reverse,start=2),
if.fuzzy.codon="solve"))
peptides_r3<-suppressWarnings(translate(subseq(dna_seq_reverse,start=3),
if.fuzzy.codon="solve"))
novel_cds_r1 <- .get_30aa_splited_seq(peptides_r1)
novel_cds_r2 <- .get_30aa_splited_seq(peptides_r2)
novel_cds_r3 <- .get_30aa_splited_seq(peptides_r3)
novel_cds_r1[,c("Strand","Frame","ID"):=list(a="-",
b="1",
c=paste(id,"-","F1",
Substring,
sep="|")),
by=.(id,Substring)]
novel_cds_r2[,c("Strand","Frame","ID"):=list(a="-",
b="2",
c=paste(id,"-","F2",
Substring,
sep="|")),
by=.(id,Substring)]
novel_cds_r3[,c("Strand","Frame","ID"):=list(a="-",
b="3",
c=paste(id,"-","F3",
Substring,
sep="|")),
by=.(id,Substring)]
all_pep<-rbindlist(list(novel_cds_f1,novel_cds_f2,novel_cds_f3,
novel_cds_r1,novel_cds_r2,novel_cds_r3))
}else
{
all_pep<-rbindlist(list(novel_cds_f1,novel_cds_f2,novel_cds_f3))
}
all_pep[,Index:=paste("NTX",cumIndex+.I,sep="")]
if(bool_get_longest){
#final_all_pep<-all_pep[,.(pep=seq[nchar(seq)==max(nchar(seq))],
#ID=ID[nchar(seq)==max(nchar(seq))]),by=.(id)]
final_all_pep<-all_pep[,.(pep=seq[nchar(seq)==max(nchar(seq))],
ID=ID[nchar(seq)==max(nchar(seq))],
Index=Index[nchar(seq)==max(nchar(seq))],
Strand=Strand[nchar(seq)==max(nchar(seq))],
Frame=Frame[nchar(seq)==max(nchar(seq))],
start=start[nchar(seq)==max(nchar(seq))],
end=end[nchar(seq)==max(nchar(seq))],
Substring=Substring[nchar(seq)==max(nchar(seq))]),
by=.(id)]
if(file.exists(outmtab)){
write.table(subset(final_all_pep,
select=c(Index,id,Strand,Frame,start,end,Substring)),
file=outmtab, sep='\t', quote=FALSE, row.names=FALSE,
append=TRUE,col.names=FALSE)
}else{
write.table(subset(final_all_pep,
select=c(Index,id,Strand,Frame,start,end,Substring)),
file=outmtab, sep='\t', quote=FALSE, row.names=FALSE)
}
#final_all_pep<-all_pep[,.(pep=seq[which.max(nchar(seq))],ID=ID[which.max(nchar(seq))]),by=.(id)]
final_all_pep[,output:=paste('>',Index,"|",ID,'\n',pep,sep=''),
by=.(ID)]
write(final_all_pep$output,file=outfa,append=TRUE)
}else{
if(file.exists(outmtab)){
write.table(subset(all_pep,
select=c(Index,id,Strand,Frame,start,end,Substring)),
file=outmtab, sep='\t', quote=FALSE, row.names=FALSE,
append=TRUE,col.names=FALSE)
}else{
write.table(subset(all_pep,
select=c(Index,id,Strand,Frame,start,end,Substring)),
file=outmtab, sep='\t', quote=FALSE, row.names=FALSE)
}
all_pep[,output:=paste('>',Index,"|",ID,'\n',seq,sep=''),by=.(ID)]
write(all_pep$output,file=outfa,append=TRUE)
}
#add var tag, add decoy sequence
xset_for<-readAAStringSet(outfa)
xset_rev<-xset_for
names(xset_for)=paste(var_tag,"|",names(xset_for),sep="")
names(xset_rev)=paste(decoy_tag,var_tag,"|",names(xset_rev),sep="")
xset_rev<-reverse(xset_rev)
outfile<-paste(outfile_name,"_txFinder.fasta",sep="")
writeXStringSet(xset_for,outfile)
if(make_decoy){
writeXStringSet(xset_rev,outfile,append=TRUE)
}
message("Write protein sequences to file: ",outfile)
return(outfile)
}
.get_30aa_splited_seq<-function(translated_seq){
df<-as.data.frame(translated_seq)
df["id"]<-rownames(df)
colnames(df)<-c("raw","id")
dt<-setDT(df)
dt_split<-dt[,.(seq=unlist(strsplit(raw, "\\*"))),by=.(id,raw)]
#must be 1L, not 1
dt_split[, cumlen := cumsum((nchar(seq)+1L)), by=list(id, raw)]
dt_split[, c("start","end"):=list(cumlen-nchar(seq),cumlen-1L),
by=list(id, raw)]
#remove the pep less than 30AA .
sub.dt_split<-subset(dt_split,(end-start)>=29)
#add column "Substring"
sub.dt_split[,Substring:=seq(1,.N),by=.(id,raw)]
sub.dt_split
}
#' @title Merge the databases from RNA-Seq and the canonical reference database
#' @description Merge the databases from RNA-Seq and the canonical reference
#' database
#' @param fa A list object containing the databases from RNA-Seq
#' @param proteinseq An object containing the canonical reference database
#' @param make_decoy A logical indicating whether to add the decoy sequences
#' @param var_tag A string will be as the prefix of sequences from RNA-Seq
#' @param decoy_tag The prefix of decoy sequences
#' @param outfile_path Output directory
#' @param outfile_name Output file name
#' @return The file name of the customized database
#' @noRd
dbcat<-function(fa,proteinseq,make_decoy=TRUE,var_tag="VAR",decoy_tag="#REV#",
outfile_path=".",outfile_name="test"){
if(is.null(fa))
{
stop("Error: fasta list not found!")
}
if(is.null(proteinseq))
{
stop("Error: \"proteinseq\" not found!")
}
## process the reference proteins
## get rid of the possible "*" (stop codon) in the end of the protein
canonical_for<-AAStringSet(x=gsub("\\*$","",proteinseq$peptide))
names(canonical_for)<-proteinseq$pro_name
canonical_rev<-canonical_for
names(canonical_rev)=paste(decoy_tag,names(canonical_for),sep="")
canonical_rev<-reverse(canonical_rev)
## process the sequences from RNA-Seq
tmp_for<-tempfile(pattern="Forward_",fileext=".fasta")
tmp_rev<-tempfile(pattern="Reverse_",fileext=".fasta")
for (i in fa)
{
xset_for<-readAAStringSet(i)
xset_rev<-xset_for
names(xset_for)=paste(var_tag,"|",names(xset_for),sep="")
names(xset_rev)=paste(decoy_tag,var_tag,"|",names(xset_rev),sep="")
xset_rev<-reverse(xset_rev)
writeXStringSet(xset_for,tmp_for,append=TRUE)
writeXStringSet(xset_rev,tmp_rev,append=TRUE)
}
xset_tmp_for<-readAAStringSet(tmp_for)
xset_tmp_rev<-readAAStringSet(tmp_rev)
outfile<-paste(outfile_path,"/",outfile_name,"_txFinder.fasta",sep="")
writeXStringSet(xset_tmp_for,outfile)
writeXStringSet(canonical_for,outfile,append=TRUE)
if(make_decoy){
writeXStringSet(xset_tmp_rev,outfile,append=TRUE)
writeXStringSet(canonical_rev,outfile,append=TRUE)
}
return(outfile)
}
#' @title Add gene nemes
#' @description Add "gene name" column to the attachments (eg. snv.tsv and idl.tsv) of report
#' if there is a demand (optional function). This function is useful for "Ensembl" IDs.
#' @param mart See detail in function \code{\link{PrepareAnnotationEnsembl}}.
#' @param report The report directory for adding gene names.
#' @return none
#' @export
#' @examples
#' \dontrun{
#' library(PGA)
#' # set the biomaRt parameters just as "PrepareAnnotationEnsembl2" did
#'
#' mart <- biomaRt::useMart("ENSEMBL_MART_ENSEMBL",
#' dataset="hsapiens_gene_ensembl",
#' host="grch37.ensembl.org",
#' path="/biomart/martservice",
#' archive=FALSE)
#' addGeneName4Ensembl(mart=mart,report="report")
#'
#' }
addGeneName4Ensembl <- function(mart, report="report"){
attributes.id <- c("ensembl_peptide_id","external_gene_id")
if(file.exists(paste(report,"files/snv.tsv",sep="/"))){
file<-paste(report,"files/snv.tsv",sep="/")
raw<-read.delim(file,header=TRUE)
protein_ids<-as.character(raw$proname)
if(length(grep("^ENS",protein_ids[1]))>0)
{
idstab <- getBM(attributes=attributes.id, mart=mart,
filters='ensembl_peptide_id', values=protein_ids)
message("........ getting HGNC gene names of SNV table\n", appendLF=FALSE)
colnames(idstab)<-c("proname","geneNameHGNC")
tb <- merge(raw, idstab, by='proname',all.x=TRUE)[,union(names(raw),names(idstab))]
write.table(tb, file=paste(report,"files/snv.tsv",sep="/"),
sep='\t', quote=FALSE,row.names=FALSE)
}else
{
message("........ No conversion is required\n", appendLF=FALSE);
}
}
if(file.exists(paste(report,"files/idl.tsv",sep="/"))){
file<-paste(report,"files/idl.tsv",sep="/")
raw<-read.delim(file,header=TRUE)
protein_ids<-as.character(raw$proname)
if(length(grep("^ENS",protein_ids[1]))>0)
{
idstab <- getBM(attributes=attributes.id, mart=mart,
filters='ensembl_peptide_id', values=protein_ids)
message("........ getting HGNC gene names of InDel table\n", appendLF=FALSE)
colnames(idstab)<-c("proname","geneNameHGNC")
tb <- merge(raw, idstab, by='proname',all.x=TRUE)[,union(names(raw),names(idstab))]
write.table(tb, file=paste(report,"files/idl.tsv",sep="/"),
sep='\t', quote=FALSE,row.names=FALSE)
}else
{
message("........ No conversion is required\n", appendLF=FALSE)
}
}
}
### See https://genome.ucsc.edu/goldenpath/help/mysql.html for how to connect
### to a MySQL server at UCSC. By default UCSC_dbselect() uses the server
### located on the US west coast.
.UCSC_dbselect <- function(dbname, from, columns=NULL, where=NULL,
server="genome-mysql.soe.ucsc.edu")
{
columns <- if (is.null(columns)) "*" else paste0(columns, collapse=",")
SQL <- sprintf("SELECT %s FROM %s", columns, from)
if (!is.null(where)) {
stopifnot(isSingleString(where))
SQL <- paste(SQL, "WHERE", where)
}
dbconn <- dbConnect(RMariaDB::MariaDB(), dbname=dbname,
username="genome",
host=server,
port=3306)
on.exit(dbDisconnect(dbconn))
dbGetQuery(dbconn, SQL)
}
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Defines functions getNovelTx .gen_splicmatrix PrepareAnnotationRefseq2 PrepareAnnotationEnsembl2 OutputVarproseq2 OutputNovelJun2 Outputaberrant2 Tab2Range
Documented in PrepareAnnotationEnsembl2 PrepareAnnotationRefseq2
## This function requires the Tab format file from Hisat2. Read Tab file contain junctions into a GRanges object.
Tab2Range <- function(tabfile)
{
options(stringsAsFactors=FALSE)
jun <- read.table(tabfile, sep='\t', header=F, stringsAsFactors=F)
#replace the strand "." with "+" and "-".
tmp<-jun[jun$V4==".",]
tmp_forward<-tmp
tmp_reverse<-tmp
tmp_forward$V4<-"+"
tmp_reverse$V4<-"-"
jun<-rbind(jun[jun$V4!=".",],tmp_forward,tmp_reverse)
part1_sta <- as.numeric(jun[,'V2'])+1-59
part1_end <- as.numeric(jun[,'V2'])+1
part2_sta <- as.numeric(jun[,'V3'])+1
part2_end <- as.numeric(jun[,'V3'])+1+59
junction <- data.frame(chr=jun[, 'V1'], id="NULL", start=jun[, 'V2'],
end=jun[,'V3'], cov=255, strand=jun$V4, part1_len=0,
part2_len=0, part1_sta, part1_end, part2_sta, part2_end)
if('chrM' %in% junction$chr) junction <- junction[-which(junction$chr=='chrM'), ]
if('MT' %in% junction$chr) junction <- junction[-which(junction$chr=='MT'), ]
junRange <- GRanges(seqnames=junction$chr, ranges=IRanges(start=junction$part1_end,
end=junction$part2_sta), strand=junction$strand, id=junction$id,
cov=junction$cov, part1_len=junction$part1_len, part2_len=junction$part2_len,
part1_sta=junction$part1_sta, part1_end=junction$part1_end,
part2_sta=junction$part2_sta, part2_end=junction$part2_end)
}
Outputaberrant2 <- function(positiontab, outfa, outmtab, coding, proteinseq,
ids, RPKM=NULL, ...){
options(stringsAsFactors=FALSE)
idx <- grep(',', positiontab[, 'varbase'], fixed=TRUE)
if(length(idx) > 0) {
muti <- positiontab[idx, ]
muti_new <- c()
for(i in 1:dim(muti)[1]){
tmp <- cbind(muti[i, 1:8],
unlist(strsplit(muti[i, 'varbase'], ',', fixed=TRUE)),
muti[i, 10])
muti_new <- rbind(muti_new, tmp)
}
colnames(muti_new) <- colnames(positiontab)
positiontab <- positiontab[-idx, ]
positiontab <- rbind(positiontab, muti_new)
}
mtable <- merge(positiontab, coding, by.x='txid', by.y='tx_id', all.x=TRUE,
stringsAsFactors = FALSE)
#mtable <- cbind(positiontab,codingseq)
mtab_plus <- subset(mtable, strand == '+')
mtab_minus <- subset(mtable, strand == '-')
str_sub(mtab_plus[, 'coding'], mtab_plus[, 'pincoding'],
nchar(mtab_plus[, 'refbase']) + mtab_plus[, 'pincoding'] - 1) <-
mtab_plus[, 'varbase']
str_sub(mtab_minus[, 'coding'], mtab_minus[, 'pincoding'],
mtab_minus[, 'pincoding'] + nchar(mtab_minus[, 'refbase']) - 1) <-
as.character(reverseComplement(DNAStringSet(mtab_minus[, 'varbase'])))
total <- rbind(mtab_plus,mtab_minus)
total<-subset(total,!is.na(coding),) # added in 20161027, some protein ids didn't have the corresponding coding sequence. Therefore, the DNAStringSet would get the error.
label <- unlist(lapply(total[,'coding'], function(x){
if(grepl ('N',x,fixed=TRUE)){
paste('with N in pos ', paste(as.character(
gregexpr('N', x, fixed=TRUE)[[1]]), collapse=' '),
sep='')
}else ""}))
total[,'coding'] <- gsub('N', 'A', total[, 'coding'], fixed=TRUE)
aa <- suppressWarnings(as.character(translate(DNAStringSet(total[, 'coding']))))
total <- cbind(total, aa, label)
pep <- apply(total, 1, function(x) unlist(strsplit(x['aa'], '\\*'))[1])
total <- cbind(total, pep)
#########remove the entry identical to normal sequence
plist <- unique(total[, 'proname'])
proteinseq <- subset(proteinseq, pro_name %in% plist)
mseq <- merge(total,proteinseq, by.x='proname', by.y='pro_name',
all=FALSE, stringsAsFactors = FALSE)
#index <- apply(mseq, 1, function(x) grep(x['pep'],x['peptide'],fixed=T))
#grep(mseq[4,'pep'],mseq[4,'peptide'])
index <-c()
for(i in 1:dim(mseq)[1]){
index <- c(index, ifelse(grep(mseq[i, 'pep'],
mseq[i, 'peptide'], fixed=TRUE) == 1, i, '' ))
#print(i)
}
if(length(index) > 0) mseq_r <- mseq[-index, ] else mseq_r <- mseq
mseq_f <- merge(mseq_r, ids, by.x='pro_name', by.y='pro_name',
all=FALSE, stringsAsFactors = FALSE)
#vartable<-cbind(Index=paste("SNV",rownames(vartable),sep=""),vartable)
mseq_f<-cbind(Index=paste("IDL",rownames(mseq_f),sep=""),mseq_f)
mseq_o<-subset(mseq_f,select=c(Index,txid,genename,txname,proname,chr,
strand,pos,refbase,varbase,pincoding,label))
write.table(mseq_o, file=outmtab, sep='\t', quote=FALSE, row.names=FALSE)
subset(mseq_f,select=c(pro_name,proname,txid,genename,txname,chr,strand,
pos,refbase,varbase,pincoding,label,gene_name,
tx_name))
outformat <- paste('>',mseq_f[,"Index"],"|", mseq_f[, 'pro_name'], "|",
mseq_f[, 'pincoding'],
" ", mseq_f[, 'refbase'], '>', mseq_f[, 'varbase'], "|",
mseq_f[, 'tx_name'], "|", mseq_f[, 'gene_name'], "|",
mseq_f[, 'description'], "|", mseq_f[, 'label'], '\n',
mseq_f[, 'pep'], sep='')
write(outformat, file=outfa)
}
OutputNovelJun2 <- function(junction_type, genome, outfa,outmtab,
#outfile_c,
proteinseq, debug=FALSE, ...){
options(stringsAsFactors=FALSE)
#ids <- subset(ids,pro_name!='')
#trans <- transcripts(txdb)
#index <- which(values(trans)[['tx_name']] %in% ids[,'tx_name'])
#pro_trans <- trans[index]
novel_junc <- subset(junction_type, jun_type != 'known junction')
if(!length(grep('chr', novel_junc[, 'seqnames'], fixed=TRUE))>0) {
novel_junc[, 'seqnames'] <- paste('chr', novel_junc[, 'seqnames'],
sep='')#
idx <- which(novel_junc[, 'seqnames'] %in% seqnames(genome))
novel_junc <- novel_junc[idx, ]#
}
###remove abnormal junctions
idx_abn <- union(which(novel_junc[, 'start'] < 0),
which(novel_junc[, 'end'] < 0))
if(length(idx_abn > 0)) novel_junc <- novel_junc[-idx_abn, ]
junRange1 <- GRanges(seqnames=novel_junc$seqnames,
ranges=IRanges(start=novel_junc$part1_sta,
end=novel_junc$part1_end),
strand=novel_junc$strand,
junction_id=novel_junc$id)
junRange2 <- GRanges(seqnames=novel_junc$seqnames,
ranges=IRanges(start=novel_junc$part2_sta,
end=novel_junc$part2_end),
strand=novel_junc$strand,
junction_id=novel_junc$id)
#match1_protx <- findOverlaps(junRange1,pro_trans)
#match2_protx <- findOverlaps(junRange2,pro_trans)
#juntransRange1 <- junRange1[unique(queryHits(match1_protx))]
#juntransRange2 <- junRange2[unique(queryHits(match2_protx))]
#junseq1 <- getSeq(genome,'chr1',start=1000,end=2000,as.character=TRUE)
###already did reverseComplement
junseq1 <- getSeq(genome, junRange1)
junseq2 <- getSeq(genome, junRange2)
junseq_cat <- DNAStringSet(mapply(function(x, y, z)
ifelse(z == '+', paste(x, y, sep=''), paste(y, x, sep='')),
as.data.frame(junseq1)[, 1],
as.data.frame(junseq2)[, 1], as.character(strand(junRange1))))#
#debug
debug=T
if(debug){
debug_file<-data.frame(seqnames=novel_junc$seqnames, part1_sta=novel_junc$part1_sta,
part1_end=novel_junc$part1_end,part2_sta=novel_junc$part2_sta,
part1_end=novel_junc$part2_end,strand=novel_junc$strand, seq1=junseq1, seq2=junseq2, seqcat=junseq_cat)
write.table(debug_file,file="debug_file.tsv",quote = F, row.names=F)
}
#index_plus <- which(strand(junRange1) == '+')
#index_minus <- which(strand(junRange1) == '-')
#seqs_plus <- junseq_cat[index_plus]
#seqs_minus <- reverseComplement(junseq_cat[index_minus])
#seqs <- c(seqs_plus, seqs_minus)
#novel_junc_new <- rbind(novel_junc[index_plus, ],
# novel_junc[index_minus, ])
novel_junc_new <- novel_junc
seqs <- junseq_cat
##Remove sequences contains NNN
Nindx <- grep('N', seqs)
if(length(Nindx) > 0){
seqs <- seqs[-Nindx]
novel_junc_new <- novel_junc_new[-Nindx, ]
}
seqs_name <- paste(paste(novel_junc_new[, 'id'], '_',
novel_junc_new[, 'seqnames'], ':',
novel_junc_new[, 'start'], '-',
novel_junc_new[, 'end'],
sep=''), novel_junc_new[, 'cov'],sep='|')#
junpepcoding <- data.frame('pro_name'=seqs_name,
'coding'=as.data.frame(seqs)[, 1])
#save(junpepcoding, file=outfile_c)
peptides_r1 <- suppressWarnings(translate(seqs))
peptides_r2 <- suppressWarnings(translate(subseq(seqs, start=2)))
peptides_r3 <- suppressWarnings(translate(subseq(seqs, start=3)))
junpos_rna_p1 <- ifelse(novel_junc_new[, 'strand'] == '+',
as.numeric(novel_junc_new[, 'part1_len']),
as.numeric(novel_junc_new[, 'part2_len'])) #
junpos_rna_p2 <- ifelse(novel_junc_new[, 'strand'] == '+',
as.numeric(novel_junc_new[, 'part1_len'])+1,
as.numeric(novel_junc_new[, 'part2_len'])+1) #
junpos_r1_p1 <- ceiling(junpos_rna_p1/3)#
junpos_r1_p2 <- ceiling(junpos_rna_p2/3) #
junpos_r2_p1 <- ceiling((junpos_rna_p1-1)/3) #
junpos_r2_p2 <- ceiling((junpos_rna_p2-1)/3)
junpos_r3_p1 <- ceiling((junpos_rna_p1-2)/3) #
junpos_r3_p2 <- ceiling((junpos_rna_p2-2)/3)
rownum<-dim(novel_junc_new)[1]
outdf<-rbind(cbind(Index=paste("JUC",1:rownum,sep=""),
frame=rep("1",rownum),
junpos_p1=junpos_r1_p1,
junpos_p2=junpos_r1_p2,
novel_junc_new,
sequence=as.data.frame(peptides_r1)[, 1]),
cbind(Index=paste("JUC",(rownum+1):(2*rownum),sep=""),
frame=rep("2",rownum),
junpos_p1=junpos_r2_p1,
junpos_p2=junpos_r2_p2,
novel_junc_new,
sequence=as.data.frame(peptides_r2)[, 1]),
cbind(Index=paste("JUC",(2*rownum+1):(3*rownum),sep=""),
frame=rep("3",rownum),
junpos_p1=junpos_r3_p1,
junpos_p2=junpos_r3_p2,
novel_junc_new,
sequence=as.data.frame(peptides_r3)[, 1]))
### remove peptide contain stop codon
index_stop <- grep('*', outdf$sequence, fixed=TRUE)
if(length(index_stop) > 0){
outdf_rmstop <- outdf[-index_stop, ]
}else outdf_rmstop <- outdf
### check if any peptides can be found in the normal database, remove those
index_nor <- lapply(outdf_rmstop$sequence, function(x)
grep(x, proteinseq[, 'peptide'], fixed=TRUE))
index_nor <- which(unlist(lapply(index_nor, length)) > 0)
if(length(index_nor) > 0){
outdf_new <- outdf_rmstop[-index_nor, ]
}else outdf_new <- outdf_rmstop
write.table(outdf_new[,-29], file=outmtab, sep='\t', quote=FALSE,
row.names=FALSE)
tmp<-paste(">",outdf_new[,'Index'],"|",outdf_new[,'seqnames'],":",
outdf_new[,"start"],"-",outdf_new[,"end"],"|ORF",
outdf_new[,"frame"]," Junpos:",outdf_new[,"junpos_p1"],"-",
outdf_new[,"junpos_p2"],"|",outdf_new[,"strand"],"|",
outdf_new[,"tx_name_part1"],"|",outdf_new[,"tx_name_part2"],"|",
outdf_new[,"jun_type"],"\n",outdf_new[,"sequence"],sep="")
write(tmp, file=outfa)
}
OutputVarproseq2 <- function(vartable, proteinseq, outfa, outmtab, ids, lablersid=FALSE,
RPKM=NULL, ...){
options(stringsAsFactors=FALSE)
abnor_index<-which(vartable$aavar=="non-synonymous")
if(length(abnor_index)!=0){
vartable<-vartable[-abnor_index,]
}
abnor_index<-which(vartable$aapos==1 & vartable$aavar=="*")
if(length(abnor_index)!=0){
vartable<-vartable[-abnor_index,]
}
vartable<-cbind(Index=paste("SNV",rownames(vartable),sep=""),vartable)
write.table(vartable, file=outmtab, sep='\t', quote=FALSE, row.names=FALSE)
nonsy <- vartable[vartable[, 'vartype'] == "non-synonymous", ]
if(lablersid){
aavar2pro <- subset(nonsy, select=c(Index,genename, txname, proname,
aaref, aapos, aavar, rsid))
}else{
aavar2pro <- subset(nonsy, select=c(Index,genename, txname, proname,
aaref, aapos, aavar))
}
aavar2pro <- aavar2pro[aavar2pro[, 'aaref']!="*", ]
#aavar2pro <- aavar2pro[aavar2pro[, 'aavar']!="*", ]
aavar2pro <- unique(aavar2pro)
plist <- unique(aavar2pro[, 'proname'])
pep <- proteinseq[proteinseq[, 'pro_name'] %in% plist, ]
pep_var <- pep
pep_all<- c()
test <- c()
for(i in 1:dim(pep_var)[1]){
#print(i)
pvar <-subset(aavar2pro,aavar2pro[,'proname'] == pep_var[i,'pro_name'])
pvar <- pvar[order(as.numeric(pvar[, 'aapos'])), ]
for(j in 1:dim(pvar)[1]){
tmp_pep_var<-pep_var[i,]
substr(tmp_pep_var[1, 'peptide'], as.integer(pvar[j, 'aapos']),
as.integer(pvar[j, 'aapos'])) <- substr(pvar[j,'aavar'],1,1)
if(tmp_pep_var[1, 'peptide']!=pep[i, 'peptide']){
if(lablersid){
var_name<-ifelse(is.null(pvar[j,'rsid']),
paste(pvar[j,"aaref"],pvar[j,"aapos"],
pvar[j,"aavar"],sep=""),
paste(pvar[j,"rsid"],":",pvar[j,"aaref"],
pvar[j,"aapos"],pvar[j,"aavar"],
sep=""))
}else{
var_name <- paste(pvar[j,"aaref"],pvar[j,"aapos"],
pvar[j,"aavar"],sep="")}
pep_name <- cbind(Index=pvar[j,"Index"],tmp_pep_var[1,],
var_name=toString(var_name))
pep_all <- rbind(pep_all, pep_name)
}else{
test <- c(test,tmp_pep_var[1, 'pro_name'])
}
}
}
#save(pep_all,file="pep_all.Rdata")
ftab <- merge(pep_all, ids, by.x='pro_name', by.y='pro_name', all=FALSE,
stringsAsFactors = FALSE)
outformat <- apply(ftab, 1, function(x)
paste('>',x['Index'],"|", x['pro_name'], "|", x['var_name'], " ",
x['tx_name.x'], "|", x['gene_name'], "|", x['description'],
'\n', unlist(strsplit(x['peptide'], '\\*'))[1], sep=''))
write(outformat, file=outfa)
}
#' @title Prepare annotation from ENSEMBL
#' @description Prepare the annotation from ENSEMBL through biomaRt. This
#' function is modified from the function \code{\link{PrepareAnnotationEnsembl}} in \pkg{customProDB}.
#' @param mart See detail in function \code{\link{PrepareAnnotationEnsembl}}.
#' @param annotation_path See detail in function \code{\link{PrepareAnnotationEnsembl}}.
#' @param splice_matrix See detail in function \code{\link{PrepareAnnotationEnsembl}}.
#' @param dbsnp See detail in function \code{\link{PrepareAnnotationEnsembl}}.
#' @param transcript_ids See detail in function \code{\link{PrepareAnnotationEnsembl}}.
#' @param COSMIC See detail in function \code{\link{PrepareAnnotationEnsembl}}.
#' @param ... Additional arguments
#' @return Several .RData file containing annotations needed for following analysis.
#' @seealso \code{\link{PrepareAnnotationRefseq2}}.
#' @export
#' @examples
#' ensembl <- biomaRt::useMart("ENSEMBL_MART_ENSEMBL", dataset="hsapiens_gene_ensembl",
#' host="grch37.ensembl.org", path="/biomart/martservice",
#' archive=FALSE)
#'
#' annotation_path <- tempdir()
#' transcript_ids <- c("ENST00000234420", "ENST00000269305", "ENST00000445888")
#'
#' PrepareAnnotationEnsembl2(mart=ensembl, annotation_path=annotation_path,
#' splice_matrix=FALSE, dbsnp=NULL, transcript_ids=transcript_ids,
#' COSMIC=FALSE)
PrepareAnnotationEnsembl2 <- function(mart, annotation_path,
splice_matrix=FALSE, dbsnp=NULL,
transcript_ids=NULL, COSMIC=FALSE,...) {
options(stringsAsFactors=FALSE)
dataset <- biomaRt:::martDataset(mart)
biomart <- biomaRt:::martBM(mart)
host <- strsplit(strsplit(biomaRt:::martHost(mart),':')[[1]][2],'//')[[1]][2]
if (!is.null(dbsnp)) {
session <- browserSession() #
if(dataset == 'hsapiens_gene_ensembl') {
if(host == 'may2009.archive.ensembl.org'){
genome(session) <- 'hg18'
dbsnps <- 'snp130'
}else{
genome(session) <- 'hg19'
dbsnps <- trackNames(session)[grep('snp', trackNames(session),
fixed=TRUE)]
}
}
if(dataset == 'mmusculus_gene_ensembl') {
if(host == 'jan2013.archive.ensembl.org' ||
host == 'oct2012.archive.ensembl.org'){
genome(session) <- 'mm10'
dbsnps <- 'snp137'
}else{
genome(session) <- 'mm9'
dbsnps <- 'snp128'
}
}
dbsnp <- pmatch(dbsnp, dbsnps)#
if (is.na(dbsnp))
stop("invalid dbsnp name for specified genome")
if (dbsnp == -1)
stop("ambiguous dbsnp name")
}
message("Prepare gene/transcript/protein id mapping information (ids.RData) ... ",
appendLF=FALSE)
if(is.null(transcript_ids)){
transcript_ids <- getBM(attributes=c("ensembl_transcript_id"),
mart=mart)[,1]
}
attributes.id <- c("ensembl_gene_id","description")
idstab <- getBM(attributes=attributes.id, mart=mart,
filters='ensembl_transcript_id', values=transcript_ids)
colnames(idstab) <- c("ensembl_gene_id","description")
attributes.tr <- c("ensembl_gene_id", "ensembl_transcript_id",
"ensembl_peptide_id")
tr <- getBM(attributes=attributes.tr, mart=mart,
filters='ensembl_transcript_id', values=transcript_ids)
colnames(tr) <- c("ensembl_gene_id", "ensembl_transcript_id",
"ensembl_peptide_id")
ids <- merge(tr, idstab, by='ensembl_gene_id')
colnames(ids) <- c('gene_name', 'tx_name', 'pro_name', 'description')
save(ids, file=paste(annotation_path, '/ids.RData', sep=''))
packageStartupMessage(" done")
message("Build TranscriptDB object (txdb.sqlite) ... ", appendLF=TRUE)
tr_coding <- subset(ids, pro_name != "")
tr_noncoding <- subset(ids, pro_name == "")
txdb<- customProDB:::makeTranscriptDbFromBiomart_archive(biomart=biomart,
dataset=dataset,
host=host,
path="/biomart/martservice",
archive=FALSE,
transcript_ids=transcript_ids)
#saveFeatures(txdb, file=paste(annotation_path,'/txdb.sqlite',sep=''))
saveDb(txdb, file=paste(annotation_path, '/txdb.sqlite', sep=''))
packageStartupMessage(" done")
#txdb_coding <- makeTranscriptDbFromBiomart_archive(biomart=biomart,
# dataset=dataset, host=host, path="/biomart/martservice", archive=FALSE,
# transcript_ids=tr_coding[, "tx_name"])
#saveFeatures(txdb_coding, file=paste(annotation_path, '/txdb_coding.sqlite', sep=''))
#saveDb(txdb_coding, file=paste(annotation_path, '/txdb_coding.sqlite', sep=''))
#txdb_noncoding <- makeTranscriptDbFromBiomart_archive(biomart=biomart,
# dataset =dataset, host=host, path="/biomart/martservice",
# archive=FALSE, transcript_ids=tr_noncoding[, "tx_name"])
#saveFeatures(txdb_noncoding, file=paste(annotation_path, '/txdb_noncoding.sqlite', sep=''))
#saveDb(txdb_noncoding, file=paste(annotation_path, '/txdb_noncoding.sqlite', sep=''))
transGrange <- transcripts(txdb)
transintxdb <- IRanges::as.data.frame(transGrange)[, c('tx_id', 'tx_name')]
#exon_anno.RData
message("Prepare exon annotation information (exon_anno.RData) ... ",
appendLF=FALSE)
attributes.exon <- c("ensembl_exon_id", "ensembl_peptide_id",
'ensembl_gene_id', "chromosome_name",
"start_position", "end_position",
"exon_chrom_start", "exon_chrom_end", "strand",
"5_utr_start", "5_utr_end", "3_utr_start",
"3_utr_end", "cds_start", "cds_end", "rank",
"ensembl_transcript_id")
# download exon information
exon <- getBM(attributes=attributes.exon, mart=mart,
filters='ensembl_transcript_id', values=transcript_ids)
colnames(exon) <- attributes.exon
exon <- merge(exon, transintxdb, by.x="ensembl_transcript_id",
by.y="tx_name")
colnames(exon) <- c("tx_name", "exon_name", "pro_name", "gene_name",
"chromosome_name", "start_position", "end_position",
"exon_chrom_start", "exon_chrom_end", "strand",
"5_utr_start", "5_utr_end", "3_utr_start",
"3_utr_end", "cds_start", "cds_end", "rank", "tx_id")
# cdsBy is a function from package "GenomicFeatures".
# It's used to extract the cds from txdb.
cdsByTx <- cdsBy(txdb, "tx", use.names=TRUE)
# convert object of IRanges to data.frame
cdss <- IRanges::as.data.frame(cdsByTx)
cds_chr_p <- data.frame(tx_name=cdss[, "group_name"],
cds_chr_start=cdss[, "start"],
cds_chr_end=cdss[, "end"],
rank=cdss[, "exon_rank"])
# substitude "group" with "group_name", modified by Shawn,"group" was just
# supported by the version before bioconductor 3.1.
cds_chr_p_coding <- subset(cds_chr_p, tx_name %in% tr_coding[, 'tx_name'])
exon <- merge(exon, cds_chr_p_coding, by.y=c("tx_name", "rank"),
by.x=c("tx_name", "rank"), all.x=TRUE)
save(exon,file=paste(annotation_path, '/exon_anno.RData', sep=''))
packageStartupMessage(" done")
# procodingseq.RData is not useful for AS construction.
message("Prepare protein coding sequence (procodingseq.RData)... ",
appendLF=FALSE)
attributes.codingseq <- c("coding", "ensembl_peptide_id",
"ensembl_transcript_id")
if(length(tr_coding[, 'pro_name']<10000)){
coding <- getBM(attributes=attributes.codingseq,
filters="ensembl_peptide_id",
values=tr_coding[, 'pro_name'],
mart=mart)
}else{
index <- floor(length(tr_coding[, 'pro_name'])/10000)
coding <- c()
for(i in 1:index) {
st <- (i-1)*10000+1
ed <- i*10000
tmp <- getBM(attributes=attributes.codingseq,
filters="ensembl_peptide_id",
values=tr_coding[st:ed, 'pro_name'], mart=mart)
coding <- rbind(coding, tmp)
#print(i)
}
tmp <- getBM(attributes=attributes.codingseq,
filters="ensembl_peptide_id",
values=tr_coding[ed+1:length(tr_coding[, 'pro_name']),
'pro_name'],
mart=mart)
coding <- rbind(coding, tmp)
}
colnames(coding) <- attributes.codingseq
tx_id <- transintxdb[match(coding[, 'ensembl_transcript_id'],
transintxdb[, 'tx_name']),
'tx_id']
procodingseq <- cbind(coding, tx_id)
colnames(procodingseq) <- c("coding", "pro_name", "tx_name", "tx_id")
save(procodingseq,file=paste(annotation_path,'/procodingseq.RData',sep=''))
packageStartupMessage(" done")
message("Prepare protein sequence (proseq.RData) ... ", appendLF=FALSE)
attributes.proseq <- c("peptide", "ensembl_peptide_id",
"ensembl_transcript_id")
if(length(tr_coding[, 'pro_name']<10000)){
proteinseq <- getBM(attributes=attributes.proseq,
filters="ensembl_peptide_id",
values=tr_coding[,'pro_name'], mart=mart)
}else{
index <- floor(length(tr_coding[, 'pro_name'])/10000)
proteinseq <- c()
for(i in 1:index) {
st <- (i-1)*10000+1
ed <- i*10000
tmp <- getBM(attributes=attributes.proseq,
filters="ensembl_peptide_id",
values= tr_coding[st:ed, 'pro_name'],
mart=mart)
proteinseq <- rbind(proteinseq, tmp)
#print(i)
}
tmp <- getBM(attributes=attributes.proseq, filters="ensembl_peptide_id",
values=tr_coding[ed+1:length(tr_coding[, 'pro_name']),
'pro_name'],
mart=mart)
proteinseq <- rbind(proteinseq, tmp)
}
colnames(proteinseq) <- c("peptide", "pro_name", "tx_name")
save(proteinseq, file=paste(annotation_path, '/proseq.RData', sep=''))
packageStartupMessage(" done")
if (!is.null(dbsnp)) {
message("Prepare dbSNP information (dbsnpinCoding.RData) ... ",
appendLF=FALSE)
if(length(dbsnps) == 1&&dbsnps == 'snp128'){
dbsnp_query <- ucscTableQuery(session, dbsnps[dbsnp],
table='snp128')
}else{
dbsnp_query <- ucscTableQuery(session, dbsnps[dbsnp],
table=paste(dbsnps[dbsnp],
'CodingDbSnp', sep=''))
}
snpCodingTab <- getTable(dbsnp_query)
snpCodingTab[, 'chrom'] <- gsub('chr', '', snpCodingTab[, 'chrom'])
chrlist <- paste(c(seq(1:22),'X','Y'))
snpCoding <- subset(snpCodingTab, chrom %in% chrlist ,
select=c(chrom:name, alleleCount, alleles))
snpCoding <- unique(snpCoding)
#snpCoding[, 'chrom'] <- gsub('chrM', 'MT', snpCoding[, 'chrom'])
#
#save(snpCoding,file=paste(annotation_path,'/snpcoding.RData',sep=''))
snpCoding <- GRanges(seqnames=snpCoding[, 'chrom'],
ranges=IRanges(start=snpCoding[, 'chromStart'],
end=snpCoding[, 'chromEnd']),
strand='*',
rsid=snpCoding[, 'name'],
alleleCount=snpCoding[, 'alleleCount'],
alleles=snpCoding[, 'alleles'])
#seqlevels(snpCoding)
#if(TRUE%in% grep('chr',seqlevels(snpCoding)) > 0 ) {
# rchar <- sub('chr','',seqlevels(snpCoding))
# names(rchar) <- seqlevels(snpCoding)
# snpCoding <- renameSeqlevels(snpCoding, rchar) }
#if('M'%in%seqlevels(snpCoding)) snpCoding <- renameSeqlevels(snpCoding, c( M='MT'))
#chrlist <- paste(c(seq(1:22),'X','Y'))
transGrange_snp <- transGrange
#transGrange_snp <- keepSeqlevels(transGrange_snp, snpCoding)
#snpCoding <- keepSeqlevels(snpCoding, transGrange_snp)
#snpCoding <- keepSeqlevels(snpCoding, transGrange)
dbsnpinCoding <- subsetByOverlaps(snpCoding,transGrange_snp)
save(dbsnpinCoding,file=paste(annotation_path, '/dbsnpinCoding.RData',
sep=''))
packageStartupMessage(" done")
}
if (COSMIC) {
message("Prepare COSMIC information (cosmic.RData) ... ",
appendLF=FALSE)
cosmic_query <- ucscTableQuery(session, 'cosmic', table='cosmic')
cosmicTab <- getTable(cosmic_query)
cosmicTab[,'chrom'] <- gsub('chrM', 'MT', cosmicTab[, 'chrom'])
cosmicTab[,'chrom'] <- gsub('chr', '', cosmicTab[, 'chrom'])
chrlist <- paste(c(seq(1:22),'X','Y','MT'))
cosmicTab <- subset(cosmicTab, chrom %in% chrlist)
cosmic <- GRanges(seqnames=cosmicTab[, 'chrom'],
ranges=IRanges(start=cosmicTab[, 'chromStart'],
end=cosmicTab[, 'chromEnd']),
strand = '*',
cosid=cosmicTab[, 'name'])
transGrange_cosmic <- transGrange
#transGrange_cosmic <- keepSeqlevels(transGrange_cosmic, cosmic)
#cosmic <- keepSeqlevels(cosmic, transGrange_cosmic)
cosmic <- subsetByOverlaps(cosmic, transGrange_cosmic)
save(cosmic,file=paste(annotation_path, '/cosmic.RData', sep=''))
packageStartupMessage(" done")
}
if(splice_matrix){
message("Prepare exon splice information (splicemax.RData) ... ",
appendLF=FALSE)
exonByTx <- exonsBy(txdb, "tx", use.names=FALSE)
index <- which(elementNROWS(exonByTx)==1)
exonByTx_mul <- exonByTx[-index]
exons_mul <- IRanges::as.data.frame(exonByTx_mul)
exonslist <- split(exons_mul, exons_mul$group)
splicemax_list <- lapply(exonslist, .gen_splicmatrix)
splicemax <- do.call(rbind, splicemax_list)
save(splicemax, file=paste(annotation_path, '/splicemax.RData', sep=''))
packageStartupMessage(" done")
}
#splice <- paste(splicemax[, 1], splicemax[, 2], sep='-')
}
#' @title Prepare annotation from Refseq
#' @description Prepare the annotation for Refseq through UCSC table browser. This
#' function is modified from the function \code{\link{PrepareAnnotationRefseq}} in \pkg{customProDB}.
#' @param genome See detail in function \code{\link{PrepareAnnotationRefseq}}.
#' @param CDSfasta See detail in function \code{\link{PrepareAnnotationRefseq}}.
#' @param pepfasta See detail in function \code{\link{PrepareAnnotationRefseq}}.
#' @param annotation_path See detail in function \code{\link{PrepareAnnotationRefseq}}.
#' @param dbsnp See detail in function \code{\link{PrepareAnnotationRefseq}}.
#' @param transcript_ids See detail in function \code{\link{PrepareAnnotationRefseq}}.
#' @param splice_matrix See detail in function \code{\link{PrepareAnnotationRefseq}}.
#' @param COSMIC See detail in function \code{\link{PrepareAnnotationRefseq}}.
#' @param ... Additional arguments
#' @return Several .RData file containing annotations needed for following analysis.
#' @seealso \code{\link{PrepareAnnotationEnsembl2}}.
#' @export
#' @examples
#' \dontrun{
#' transcript_ids <- c("NM_001126112", "NM_033360", "NR_073499")
#' pepfasta <- system.file("extdata", "refseq_pro_seq.fasta",
#' package="customProDB")
#' CDSfasta <- system.file("extdata", "refseq_coding_seq.fasta",
#' package="customProDB")
#' annotation_path <- tempdir()
#' PrepareAnnotationRefseq2(genome='hg19', CDSfasta, pepfasta, annotation_path,
#' dbsnp=NULL, transcript_ids=transcript_ids,
#' splice_matrix=FALSE, COSMIC=FALSE)
#' }
PrepareAnnotationRefseq2 <- function(genome='hg19', CDSfasta, pepfasta,
annotation_path, dbsnp=NULL,
transcript_ids=NULL, splice_matrix=FALSE,
COSMIC=FALSE, ...) {
options(stringsAsFactors=FALSE)
session <- browserSession()
genome(session) <- genome
tablename <- 'refGene'
message("Build TranscriptDB object (txdb.sqlite) ... ", appendLF=TRUE)
txdb <- makeTxDbFromUCSC(genome=genome, tablename=tablename,
transcript_ids=transcript_ids)
saveDb(txdb, file=paste(annotation_path, '/txdb.sqlite', sep=''))
packageStartupMessage(" done")
message("Prepare gene/transcript/protein id mapping information (ids.RData) ... ",
appendLF=FALSE)
query_refGene <- ucscTableQuery(session, "refSeqComposite", table="refGene")
refGene <- getTable(query_refGene)
if(!is.null(transcript_ids))
{
refGene <- subset(refGene, name %in% transcript_ids)
}
reflink <- .UCSC_dbselect("hgFixed", "refLink")
ids <- subset(reflink, mrnaAcc %in% refGene[, 'name'], select = name:protAcc)
colnames(ids) <- c('gene_name', 'description', 'tx_name', 'pro_name')
save(ids, file=paste(annotation_path, '/ids.RData', sep=''))
packageStartupMessage(" done")
#tr_coding <- subset(ids,pro_name!="")
#tr_noncoding <- subset(ids,pro_name == "")
#txdb_coding <- makeTranscriptDbFromUCSC(genome=genome, tablename=tablename,
# transcript_ids=tr_coding[, "tx_name"] )
#saveDb(txdb_coding,
# file=paste(annotation_path, '/txdb_coding.sqlite', sep=''))
#txdb_noncoding <- makeTranscriptDbFromUCSC(genome=genome,
# tablename=tablename, transcript_ids=tr_noncoding[, "tx_name"] )
#saveDb(txdb_noncoding,
# file=paste(annotation_path, '/txdb_noncoding.sqlite', sep=''))
message("Prepare exon annotation information (exon_anno.RData) ... ",
appendLF=FALSE)
transGrange <- transcripts(txdb)
tr <- IRanges::as.data.frame(transGrange)
cdsByTx <- cdsBy(txdb, "tx", use.names=FALSE)
exonByTx <- exonsBy(txdb, "tx", use.names=FALSE)
fiveutrByTx <- fiveUTRsByTranscript(txdb, use.names=FALSE)
threeutrByTx <- threeUTRsByTranscript(txdb, use.names=FALSE)
#####################################################
# get error in most recent version of IRanges package
# previous: rownames after unlist 1.1 1.2 1.3
# now: .1 .2 .3 ... were removed, so there are some rows with same rownames
######################################################
#cdss <- IRanges::as.data.frame(IRanges::unlist(cdsByTx))
#exons <- IRanges::as.data.frame(IRanges::unlist(exonByTx))
#fiveutrs <- IRanges::as.data.frame(IRanges::unlist(fiveutrByTx))
#threeutrs <- IRanges::as.data.frame(IRanges::unlist(threeutrByTx))
cdss <- IRanges::as.data.frame(cdsByTx)
exons <- IRanges::as.data.frame(exonByTx)
fiveutrs <- IRanges::as.data.frame(fiveutrByTx)
threeutrs <- IRanges::as.data.frame(threeutrByTx)
#txid <- matrix(unlist(strsplit(rownames(exons), '\\.')), ncol = 2,
#byrow =T)[, 1]
#txid <- gsub('=','\\.', txid)
#substitude "group" with "group_name", modified by Shawn."group" was just
#supported by the version before bioconductor 3.1
exon_p <- data.frame(txid=exons[, "group_name"], chr=exons[, "seqnames"],
exon_s=exons[, "start"], exon_e=exons[, "end"],
exon_rank=exons[, "exon_rank"])
exon2tr <- merge(exon_p, tr,by.y="tx_id", by.x="txid")
exon2tr <- exon2tr[, -which(names(exon2tr) %in% c("seqnames", "width"))]
#txid <- matrix(unlist(strsplit(rownames(cdss), '\\.')), ncol = 2,
# byrow =T)[, 1]
#txid <- gsub('=','\\.',txid)
#substitude "group" with "group_name", modified by Shawn
cds_p <- data.frame(txid=cdss[, "group_name"], cds_s=cdss[, "start"],
cds_e=cdss[, "end"], exon_rank=cdss[, "exon_rank"],
width=cdss[, "width"])
ttt <- split(cds_p, cds_p$txid)
cds_p_new_list <-lapply(ttt, function(x){
#len <- x[,'cds_e']-x[,'cds_s']+1
#cum <- cumsum(len)
cum <- cumsum(x[, 'width'])
rdis <- cbind(c(1, cum[1:length(cum)-1]+1), cum)
colnames(rdis) <- c('cds_start', 'cds_end')
tmp <- cbind(x, rdis)
tmp
})
cds_p_new <- do.call(rbind, cds_p_new_list)
cds_p_new <- cds_p_new[, -which(names(cds_p_new) %in% c("width"))]
#for(i in 1:length(ttt)) {
# print(i)
# ttt1 <- ttt[[i]]
# len <- ttt1[,'cds_e']-ttt1[,'cds_s']+1
# cum <- cumsum(len)
# rdis <- cbind(c(1,cum[1:length(cum)-1]+1),cum)
# colnames(rdis) <- c('cds_start','cds_end')
# tmp <- cbind(ttt1,rdis)
# cds_p_new <- rbind(cds_p_new,tmp)
#}
cds2exon <- merge(exon2tr, cds_p_new, by.x=c("txid", "exon_rank"),
by.y=c("txid", "exon_rank"), all.x = TRUE)
#txid <- matrix(unlist(strsplit(rownames(fiveutrs), '\\.')), ncol = 2,
# byrow =T)[, 1]
#txid <- gsub('=','\\.', txid)
#substitude "group" with "group_name", modified by Shawn
fiveutr_p <- data.frame(txid=fiveutrs[, "group_name"],
fiveutr_s=fiveutrs[, "start"],
fiveutr_e=fiveutrs[, "end"],
exon_rank=fiveutrs[, "exon_rank"])
fiveutr2exon <- merge(cds2exon, fiveutr_p, by.x=c("txid", "exon_rank"),
by.y =c("txid", "exon_rank"), all.x = TRUE)
#txid <- matrix(unlist(strsplit(rownames(threeutrs),'\\.')),
#ncol = 2,byrow =T)[, 1]
#txid <- gsub('=','\\.', txid)
#substitude "group" with "group_name", modified by Shawn
threeutr_p <- data.frame(txid=threeutrs[, "group_name"],
threeutr_s=threeutrs[, "start"],
threeutr_e=threeutrs[, "end"],
exon_rank=threeutrs[, "exon_rank"])
threeutr2exon <- merge(fiveutr2exon, threeutr_p,
by.x=c("txid", "exon_rank"),
by.y=c("txid", "exon_rank"), all.x = TRUE)
#exon <- merge(threeutr2exon,ids,by.x=c("tx_name"),by.y="tx_name",
#all.x = TRUE)
exon <- threeutr2exon[order(threeutr2exon$txid, threeutr2exon$exon_rank), ]
colnames(exon) <- c("tx_id","rank", "chromosome_name", "exon_chrom_start",
"exon_chrom_end", "start_position", "end_position",
"strand", "tx_name", "cds_chr_start", "cds_chr_end",
"cds_start", "cds_end", "5_utr_start",
"5_utr_end", "3_utr_start", "3_utr_end")
pro_name <- ids[match(exon[, 'tx_name'], ids[, 'tx_name']), 'pro_name']
gene_name <- ids[match(exon[, 'tx_name'], ids[, 'tx_name']), 'gene_name']
exon <- cbind(exon, pro_name, gene_name)
save(exon, file=paste(annotation_path, '/exon_anno.RData', sep=''))
packageStartupMessage(" done")
message("Prepare protein sequence (proseq.RData) ... ", appendLF=FALSE)
pro_seqs <- readAAStringSet(pepfasta, format= 'fasta')
pro_name_v <- names(pro_seqs)
pro_name <- unlist(lapply(pro_name_v, function(x) strsplit(x, '\\.')[[1]][1]))
tx_name <- ids[match(pro_name, ids[, 'pro_name']), 'tx_name']
proteinseq <- as.data.frame(pro_seqs)
proteinseq <- cbind(proteinseq, pro_name_v, pro_name, tx_name)
colnames(proteinseq) <- c("peptide", "pro_name_v", "pro_name", "tx_name")
proteinseq <- subset(proteinseq, tx_name %in% refGene[, 'name'])
save(proteinseq, file=paste(annotation_path, '/proseq.RData', sep=''))
packageStartupMessage(" done")
message("Prepare protein coding sequence (procodingseq.RData)... ",
appendLF=FALSE)
cds_seqs <- readDNAStringSet(CDSfasta, format= 'fasta')
tx_name_tmp <- unlist(lapply(names(cds_seqs), function(x)
strsplit(x, ' ')[[1]][1]))
tx_name <- unlist(lapply(tx_name_tmp, function(x)
paste(strsplit(x, '_')[[1]][3:4], collapse='_')))
tx_range_tmp <- unlist(lapply(names(cds_seqs), function(x)
strsplit(x, ' ')[[1]][2]))
tx_chr <- unlist(lapply(tx_range_tmp, function(x)
substring(strsplit(x, ':')[[1]][1],7)))
tx_cds_sta <- unlist(lapply(tx_range_tmp, function(x)
strsplit(strsplit(x, ':')[[1]][2], '-')[[1]][[1]]))
tx_cds_end <- unlist(lapply(tx_range_tmp, function(x)
strsplit(strsplit(x, ':')[[1]][2], '-')[[1]][[2]]))
tx_name_cds <- refGene[match(paste(tx_name, tx_chr, tx_cds_sta, tx_cds_end,
sep=' '),
paste(refGene[, 'name'], refGene[, 'chrom'],
refGene[, 'cdsStart']+1,
refGene[, 'cdsEnd'], sep=' ')),
c('name','chrom','txStart','txEnd')]
tx_id <- tr[match(paste(tx_name_cds[, 'name'], tx_name_cds[, 'chrom'],
tx_name_cds[, 'txStart']+1,
tx_name_cds[, 'txEnd'],
sep=' '),
paste(tr[, 'tx_name'], tr[, 'seqnames'], tr[, 'start'],
tr[, 'end'], sep=' ')), 'tx_id']
pro_name <- ids[match(tx_name,ids[, 'tx_name']), 'pro_name']
procodingseq <- as.data.frame(cds_seqs)
procodingseq <- cbind(procodingseq, names(cds_seqs), pro_name,
tx_name, tx_id)
colnames(procodingseq) <- c("coding", "tx_name_full", "pro_name",
"tx_name", "tx_id")
procodingseq <- subset(procodingseq, tx_name %in% refGene[, 'name'])
save(procodingseq, file=paste(annotation_path, '/procodingseq.RData',
sep=''))
packageStartupMessage(" done")
if (!is.null(dbsnp)) {
message("Prepare dbSNP information (dbsnpinCoding.RData) ... ",
appendLF=FALSE)
dbsnps <- trackNames(session)[grep('snp', trackNames(session),
fixed=TRUE)]
dbsnp <- pmatch(dbsnp, dbsnps)
if (is.na(dbsnp))
stop("invalid dbsnp name for specified genome")
if (dbsnp == -1)
stop("ambiguous dbsnp name")
dbsnp_query <- ucscTableQuery(session, dbsnps[dbsnp],
table=paste(dbsnps[dbsnp],
'CodingDbSnp', sep=''))
snpCodingTab <- getTable(dbsnp_query)
snpCoding <- subset(snpCodingTab,transcript %in% refGene[, 'name'],
select=c(chrom:name, alleleCount, alleles))
snpCoding <- unique(snpCoding)
#save(snpCoding,file=paste(annotation_path, '/snpcoding.RData', sep=''))
dbsnpinCoding <- GRanges(seqnames=snpCoding[, 'chrom'],
ranges=IRanges(start=snpCoding[, 'chromStart'],
end=snpCoding[, 'chromEnd']),
strand='*',
rsid=snpCoding[, 'name'],
alleleCount=snpCoding[, 'alleleCount'],
alleles=snpCoding[, 'alleles'])
save(dbsnpinCoding, file=paste(annotation_path, '/dbsnpinCoding.RData',
sep=''))
packageStartupMessage(" done")
}
if (COSMIC) {
#cosmic <- trackNames(session)[grep('cosmic',trackNames(session),
#fixed=T)]
message("Prepare COSMIC information (cosmic.RData) ...",appendLF=FALSE)
cosmic_query <- ucscTableQuery(session, 'cosmic', table='cosmic')
cosmicTab <- getTable(cosmic_query)
cosmic <- GRanges(seqnames=cosmicTab[, 'chrom'],
ranges=IRanges(start=cosmicTab[, 'chromStart'],
end=cosmicTab[, 'chromEnd']),
strand = '*', cosid=cosmicTab[,'name'])
#cosmic <- keepSeqlevels(cosmic,transGrange)
cosmic <- subsetByOverlaps(cosmic, transGrange)
save(cosmic,file=paste(annotation_path, '/cosmic.RData', sep=''))
packageStartupMessage(" done")
}
if(splice_matrix){
message("Prepare exon splice information (splicemax.RData) ... ",
appendLF=FALSE)
index <- which(elementNROWS(exonByTx)==1)
exonByTx_mul <- exonByTx[-index]
exons_mul <- IRanges::as.data.frame(exonByTx_mul)
exonslist <- split(exons_mul, exons_mul$group)
#system.time( exonByTx <- exonsBy(txdb,"tx", use.names=F))
splicemax_list <- lapply(exonslist, .gen_splicmatrix)
splicemax <- do.call(rbind, splicemax_list)
save(splicemax, file=paste(annotation_path, '/splicemax.RData',
sep=''))
packageStartupMessage(" done")
}
}
.gen_splicmatrix <- function(x, ...) {
mystrand=x[1,'strand']
a=x[,'exon_rank']
b=x[,'exon_id']
n=length(a)
if (mystrand=='+'){
tmp=order(a)
}else{
tmp=order(a,decreasing=TRUE)
}
mm=cbind(b[tmp[1:(n-1)]], b[tmp[2:n]])
return(mm)
}
##' @title Create protein database for novel transcripts
##' @description Create protein database for novel transcripts
##' @param gffFile A GFF format file containing novel transcripts information
##' @param outmtab A txt format file containing the novel transcripts information
##' @param outfa The output fasta format protein sequence file
##' @param outgtf The output GTF format file
##' @param bool_get_longest When it's set as TRUE, the longest sequences will be
##' retained after the DNA sequences are six-frame translated into protein
##' sequences. Otherwise, the protein sequences more than 30 aa are retained.
##' @param organism What is the Genus and species of this organism.Please use
##' proper scientific nomenclature for example: "Homo sapiens" and not "human",
##' default is "Homo sapiens".
##' @param genome Genome information
##' @param ... Additional arguments
##' @return The database file(s)
##' @noRd
getNovelTx=function(gffFile="transcripts.gtf",
outmtab="novel_transcripts_ntx.tab",
outfa="novel_transcripts_ntx.fasta",
outgtf="novel_transcripts_ntx.gtf",
bool_get_longest=TRUE,genome=NULL,organism="Homo sapiens"){
options(stringsAsFactors = FALSE)
if(is.null(genome)){
stop("genome not found!")
}
########## Params
if(file.exists(outfa)){
file.remove(outfa)
}
tmpdir<-tempdir()
chromLen<-seqlengths(genome)
#######filter
gff <- import(gffFile)
novel_gff<-gff[substr(gff$transcript_id,1,5) =="CUFF."]
#justify the chromosomes are ucsc format or ensembl format
if(any(c(1:22, "X", "Y") %in% seqlevels(novel_gff))){
# restricting this extraction to the 24 chromosomes
seqlevels(novel_gff, pruning.mode="coarse") <- c(1:22, "X", "Y")
seqlevels(novel_gff) <- paste0("chr", seqlevels(novel_gff)) # rename
}else{
seqlevels(novel_gff, pruning.mode="coarse") <-
paste("chr",c(1:22, "X", "Y"), sep="")
}
novel_tx<-novel_gff[novel_gff$type=="transcript"]
#record the strand info
#dict_strand<-dlply(as.data.frame(novel_tx),.(transcript_id),
# summarize,strand=strand)
vector_nonstrand <- novel_tx[strand(novel_tx)=="*"]$transcript_id
vector_forward <- novel_tx[strand(novel_tx)=="+"]$transcript_id
vector_reverse <- novel_tx[strand(novel_tx)=="-"]$transcript_id
#the coordinate info can be found here.
export(novel_gff, outgtf, format="gtf")
strand(novel_gff)<-"+" #change the strand
tmpgff<-paste(tmpdir,"novel_transcripts_strand_forward.gtf",sep="/")
export(novel_gff, tmpgff, format="gtf")
#cat(paste("temp dir:",tmpdir,""sep=""))
#chrominfo<-data.frame(names(chromLen),as.vector(chromLen),
# rep(FALSE,length(chromLen)))
#colnames(chrominfo)<-c("chrom","length","is_circular")
# makeTxDbFromGFF {GenomicFeatures}
# Make a TxDb object from annotations available as a GFF3 or GTF file
#txdb <- makeTxDbFromGFF(file=tmpgff,format="gtf",
# exonRankAttributeName="exon_number",
# species=species,
# circ_seqs=names(chromLen))
txdb <- makeTxDbFromGFF(file=tmpgff,format="gtf",
#exonRankAttributeName="exon_number",
organism=organism)
#if(interactive()) {
#saveDb(txdb,file="TESTGTF.sqlite")
#}
## Extract the exon ranges grouped by transcript from 'txdb':
transcripts <- exonsBy(txdb, by="tx", use.names=TRUE)
#justify the chromosomes are ucsc format or ensembl format
#if(any(c(1:22, "X", "Y") %in% seqlevels(transcripts)))
#{
# restricting this extraction to the 24 chromosomes
#seqlevels(transcripts, pruning.mode="coarse") <- c(1:22, "X", "Y")
#seqlevels(transcripts) <- paste0("chr", seqlevels(transcripts)) # rename
#}
## Extract the transcript sequences from the genome:
tx_seqs <- extractTranscriptSeqs(genome, transcripts)
cumIndex <-0
if(length(vector_reverse)>0)
{
message("====== Processing reverse strand ======")
tx_seqs_reverse<-tx_seqs[match(vector_reverse,names(tx_seqs))]
tx_seqs_normal<-reverseComplement(tx_seqs_reverse)
peptides_r1<-suppressWarnings(translate(tx_seqs_normal,
if.fuzzy.codon="solve"))
peptides_r2<-suppressWarnings(translate(subseq(tx_seqs_normal,start=2),
if.fuzzy.codon="solve"))
peptides_r3<-suppressWarnings(translate(subseq(tx_seqs_normal,start=3),
if.fuzzy.codon="solve"))
#tmp_vector<-unlist(strsplit(as.data.frame(peptides_r1)$x, "\\*"))
#way1: find the longest sequence
#tmp_vector[str_length(tmp_vector)==max(str_length(tmp_vector))]
#way2: find all of the sequences more than 30 AA
#tmp_vector[str_length(tmp_vector)>=30]
novel_cds_r1 <- .get_30aa_splited_seq(peptides_r1)
novel_cds_r2 <- .get_30aa_splited_seq(peptides_r2)
novel_cds_r3 <- .get_30aa_splited_seq(peptides_r3)
novel_cds_r1[,c("Strand","Frame","ID"):=list(a="-",
b="1",
c=paste(id,"-","F1",
Substring,
sep="|")),
by=.(id,Substring)]
novel_cds_r2[,c("Strand","Frame","ID"):=list(a="-",
b="2",
c=paste(id,"-","F2",
Substring,
sep="|")),
by=.(id,Substring)]
novel_cds_r3[,c("Strand","Frame","ID"):=list(a="-",
b="3",
c=paste(id,"-","F3",
Substring,
sep="|")),
by=.(id,Substring)]
all_pep<-rbindlist(list(novel_cds_r1,novel_cds_r2,novel_cds_r3))
all_pep[,Index:=paste("NTX",cumIndex+.I,sep="")]
cumIndex<-dim(all_pep)[1]+cumIndex;
#all_pep<-rbind(cbind(names_r1,novel_cds_r1[,"subpep.seq"]),
#cbind(names_r2,novel_cds_r2[,"subpep.seq"]),
#cbind(names_r3,novel_cds_r3[,"subpep.seq"]))
if(bool_get_longest){
final_all_pep<-all_pep[,.(pep=seq[nchar(seq)==max(nchar(seq))],
ID=ID[nchar(seq)==max(nchar(seq))],
Index=Index[nchar(seq)==max(nchar(seq))],
Strand=Strand[nchar(seq)==max(nchar(seq))],
Frame=Frame[nchar(seq)==max(nchar(seq))],
start=start[nchar(seq)==max(nchar(seq))],
end=end[nchar(seq)==max(nchar(seq))],
Substring=Substring[nchar(seq)==max(nchar(seq))]),
by=.(id)]
#final_all_pep<-all_pep[,.(pep=seq[which.max(nchar(seq))],
#ID=ID[which.max(nchar(seq))]),by=.(id)]
write.table(subset(final_all_pep,
select=c(Index,id,Strand,Frame,start,end,Substring)),
file=outmtab, sep='\t', quote=FALSE, row.names=FALSE)
final_all_pep[,output:=paste('>',Index,"|",ID,'\n',pep,sep=''),
by=.(ID)]
write(final_all_pep$output,file=outfa,append=TRUE)
}else{
write.table(subset(all_pep,
select=c(Index,id,Strand,Frame,start,end,Substring)),
file=outmtab, sep='\t', quote=FALSE, row.names=FALSE)
all_pep[,output:=paste('>',Index,"|",ID,'\n',seq,sep=''),by=.(ID)]
write(all_pep$output,file=outfa,append=TRUE)
}
}
if(length(vector_forward)>0)
{
message("====== Processing forward strand ======")
tx_seqs_forward<-tx_seqs[match(vector_forward,names(tx_seqs))]
tx_seqs_normal<-tx_seqs_forward
peptides_f1<-suppressWarnings(translate(tx_seqs_normal,
if.fuzzy.codon="solve"))
peptides_f2<-suppressWarnings(translate(subseq(tx_seqs_normal,start=2),
if.fuzzy.codon="solve"))
peptides_f3<-suppressWarnings(translate(subseq(tx_seqs_normal,start=3),
if.fuzzy.codon="solve"))
#tmp_vector<-unlist(strsplit(as.data.frame(peptides_f1)$x, "\\*"))
#way1: find the longest sequence
#tmp_vector[str_length(tmp_vector)==max(str_length(tmp_vector))]
#way2: find all of the sequences more than 30 AA
#tmp_vector[str_length(tmp_vector)>=30]
novel_cds_f1 <- .get_30aa_splited_seq(peptides_f1)
novel_cds_f2 <- .get_30aa_splited_seq(peptides_f2)
novel_cds_f3 <- .get_30aa_splited_seq(peptides_f3)
novel_cds_f1[,c("Strand","Frame","ID"):=list(a="+",
b="1",
c=paste(id,"+","F1",
Substring,
sep="|")),
by=.(id,Substring)]
novel_cds_f2[,c("Strand","Frame","ID"):=list(a="+",
b="2",
c=paste(id,"+","F2",
Substring,
sep="|")),
by=.(id,Substring)]
novel_cds_f3[,c("Strand","Frame","ID"):=list(a="+",
b="3",
c=paste(id,"+","F3",
Substring,
sep="|")),
by=.(id,Substring)]
all_pep<-rbindlist(list(novel_cds_f1,novel_cds_f2,novel_cds_f3))
#all_pep<-rbind(cbind(names_f1,novel_cds_f1[,"subpep.seq"]),
#cbind(names_f2,novel_cds_f2[,"subpep.seq"]),
#cbind(names_f3,novel_cds_f3[,"subpep.seq"]))
all_pep[,Index:=paste("NTX",cumIndex+.I,sep="")]
cumIndex<-dim(all_pep)[1]+cumIndex;
if(bool_get_longest){
#final_all_pep<-all_pep[,.(pep=seq[which.max(nchar(seq))],
#ID=ID[which.max(nchar(seq))]),by=.(id)]
final_all_pep<-all_pep[,.(pep=seq[nchar(seq)==max(nchar(seq))],
ID=ID[nchar(seq)==max(nchar(seq))],
Index=Index[nchar(seq)==max(nchar(seq))],
Strand=Strand[nchar(seq)==max(nchar(seq))],
Frame=Frame[nchar(seq)==max(nchar(seq))],
start=start[nchar(seq)==max(nchar(seq))],
end=end[nchar(seq)==max(nchar(seq))],
Substring=Substring[nchar(seq)==max(nchar(seq))]),
by=.(id)]
if(file.exists(outfa)){
write.table(subset(final_all_pep,
select=c(Index,id,Strand,Frame,start,end,Substring)),
file=outmtab, sep='\t', quote=FALSE, row.names=FALSE,
append=TRUE,col.names=FALSE)
}else{
write.table(subset(final_all_pep,
select=c(Index,id,Strand,Frame,start,end,Substring)),
file=outmtab, sep='\t', quote=FALSE, row.names=FALSE)
}
final_all_pep[,output:=paste('>',Index,"|",ID,'\n',pep,sep=''),
by=.(ID)]
write(final_all_pep$output,file=outfa,append=TRUE)
}else{
if(file.exists(outfa)){
write.table(subset(all_pep,
select=c(Index,id,Strand,Frame,start,end,Substring)),
file=outmtab, sep='\t', quote=FALSE, row.names=FALSE,
append=TRUE,col.names=FALSE)
}else{
write.table(subset(all_pep,
select=c(Index,id,Strand,Frame,start,end,Substring)),
file=outmtab, sep='\t', quote=FALSE, row.names=FALSE)
}
all_pep[,output:=paste('>',Index,"|",ID,'\n',seq,sep=''),by=.(ID)]
write(all_pep$output,file=outfa,append=TRUE)
}
}
if(length(vector_nonstrand)>0){
message("====== Processing unknown strand ======")
tx_seqs_non<-tx_seqs[match(vector_nonstrand,names(tx_seqs))]
tx_seqs_normal<-tx_seqs_non
peptides_f1<-suppressWarnings(translate(tx_seqs_normal,
if.fuzzy.codon="solve"))
peptides_f2<-suppressWarnings(translate(subseq(tx_seqs_normal,start=2),
if.fuzzy.codon="solve"))
peptides_f3<-suppressWarnings(translate(subseq(tx_seqs_normal,start=3),
if.fuzzy.codon="solve"))
#tmp_vector<-unlist(strsplit(as.data.frame(peptides_f1)$x, "\\*"))
#way1: find the longest sequence
#tmp_vector[str_length(tmp_vector)==max(str_length(tmp_vector))]
#way2: find all of the sequences more than 30 AA
#tmp_vector[str_length(tmp_vector)>=30]
novel_cds_f1 <- .get_30aa_splited_seq(peptides_f1)
novel_cds_f2 <- .get_30aa_splited_seq(peptides_f2)
novel_cds_f3 <- .get_30aa_splited_seq(peptides_f3)
novel_cds_f1[,c("Strand","Frame","ID"):=list(a="+",
b="1",
c=paste(id,"+","F1",
Substring,
sep="|")),
by=.(id,Substring)]
novel_cds_f2[,c("Strand","Frame","ID"):=list(a="+",
b="2",
c=paste(id,"+","F2",
Substring,
sep="|")),
by=.(id,Substring)]
novel_cds_f3[,c("Strand","Frame","ID"):=list(a="+",
b="3",
c=paste(id,"+","F3",
Substring,
sep="|")),
by=.(id,Substring)]
############ reverseComplement
tx_seqs_normal<-reverseComplement(tx_seqs_non)
peptides_r1<-suppressWarnings(translate(tx_seqs_normal,
if.fuzzy.codon="solve"))
peptides_r2<-suppressWarnings(translate(subseq(tx_seqs_normal,start=2),
if.fuzzy.codon="solve"))
peptides_r3<-suppressWarnings(translate(subseq(tx_seqs_normal,start=3),
if.fuzzy.codon="solve"))
#tmp_vector<-unlist(strsplit(as.data.frame(peptides_r1)$x, "\\*"))
#way1: find the longest sequence
#tmp_vector[str_length(tmp_vector)==max(str_length(tmp_vector))]
#way2: find all of the sequences more than 30 AA
#tmp_vector[str_length(tmp_vector)>=30]
novel_cds_r1 <- .get_30aa_splited_seq(peptides_r1)
novel_cds_r2 <- .get_30aa_splited_seq(peptides_r2)
novel_cds_r3 <- .get_30aa_splited_seq(peptides_r3)
novel_cds_r1[,c("Strand","Frame","ID"):=list(a="-",
b="1",
c=paste(id,"-","F1",
Substring,
sep="|")),
by=.(id,Substring)]
novel_cds_r2[,c("Strand","Frame","ID"):=list(a="-",
b="2",
c=paste(id,"-","F2",
Substring,
sep="|")),
by=.(id,Substring)]
novel_cds_r3[,c("Strand","Frame","ID"):=list(a="-",
b="3",
c=paste(id,"-","F3",
Substring,
sep="|")),
by=.(id,Substring)]
all_pep<-rbindlist(list(novel_cds_f1,novel_cds_f2,novel_cds_f3,
novel_cds_r1,novel_cds_r2,novel_cds_r3))
all_pep[,Index:=paste("NTX",cumIndex+.I,sep="")]
if(bool_get_longest){
#final_all_pep<-all_pep[,.(pep=seq[nchar(seq)==max(nchar(seq))],
#ID=ID[nchar(seq)==max(nchar(seq))]),by=.(id)]
final_all_pep<-all_pep[,.(pep=seq[nchar(seq)==max(nchar(seq))],
ID=ID[nchar(seq)==max(nchar(seq))],
Index=Index[nchar(seq)==max(nchar(seq))],
Strand=Strand[nchar(seq)==max(nchar(seq))],
Frame=Frame[nchar(seq)==max(nchar(seq))],
start=start[nchar(seq)==max(nchar(seq))],
end=end[nchar(seq)==max(nchar(seq))],
Substring=Substring[nchar(seq)==max(nchar(seq))]),
by=.(id)]
if(file.exists(outmtab)){
write.table(subset(final_all_pep,
select=c(Index,id,Strand,Frame,start,end,Substring)),
file=outmtab, sep='\t', quote=FALSE, row.names=FALSE,
append=TRUE,col.names=FALSE)
}else{
write.table(subset(final_all_pep,
select=c(Index,id,Strand,Frame,start,end,Substring)),
file=outmtab, sep='\t', quote=FALSE, row.names=FALSE)
}
#final_all_pep<-all_pep[,.(pep=seq[which.max(nchar(seq))],ID=ID[which.max(nchar(seq))]),by=.(id)]
final_all_pep[,output:=paste('>',Index,"|",ID,'\n',pep,sep=''),
by=.(ID)]
write(final_all_pep$output,file=outfa,append=TRUE)
}else{
if(file.exists(outmtab)){
write.table(subset(all_pep,
select=c(Index,id,Strand,Frame,start,end,Substring)),
file=outmtab, sep='\t', quote=FALSE, row.names=FALSE,
append=TRUE,col.names=FALSE)
}else{
write.table(subset(all_pep,
select=c(Index,id,Strand,Frame,start,end,Substring)),
file=outmtab, sep='\t', quote=FALSE, row.names=FALSE)
}
all_pep[,output:=paste('>',Index,"|",ID,'\n',seq,sep=''),by=.(ID)]
write(all_pep$output,file=outfa,append=TRUE)
}
}
}
#' @title Create protein database based on the transcripts from de novo
#' reconstruction of transcriptome from RNA-seq data
#' @description Create protein database based on the transcripts from de novo
#' reconstruction of transcriptome from RNA-seq data.
#' @param infa A FASTA format file containing transcript sequences
#' (such as derived from de novo transcriptome assembly by Trinity)
#' @param bool_use_3frame A logical variable indicating whether to translate the
#' raw sequences with 3-frame (forward). Default is 6-frame translation (FALSE).
#' @param outmtab A txt format file containing the novel transcripts information
#' @param outfa The output fasta format protein sequence file
#' @param bool_get_longest When it's set as TRUE, only the longest protein sequences will be
#' retained after the raw sequences are translated with 3-frame or 6-frame.
#' Otherwise, all the protein sequences with longer than 30 aa will be retained.
#' @param make_decoy A logical variable indicating whether to add the decoy sequences.
#' @param decoy_tag The prefix of decoy sequence IDs.
#' @param outfile_name Output file name
#' @return The database file(s)
#' @export
#' @examples
#' transcript_seq_file <- system.file("extdata/input", "Trinity.fasta",package="PGA")
#' createProDB4DenovoRNASeq(infa=transcript_seq_file)
createProDB4DenovoRNASeq<-function(infa="./trinity.fasta",
bool_use_3frame=FALSE,
outmtab="novel_transcripts_ntx.tab",
outfa="./novel_transcripts_ntx.fasta",
bool_get_longest=TRUE,
make_decoy=TRUE,
decoy_tag="#REV#",
outfile_name="test"){
options(stringsAsFactors=FALSE)
if(file.exists(outfa)){
file.remove(outfa)
}
var_tag <- "DNO"
cumIndex <-0
dna_seq_forward<-readDNAStringSet(infa)
names(dna_seq_forward)<-gsub(" .+","",names(dna_seq_forward)) #get the id, remove the description
message("....... Proteomic database construction ! ............")
# forward strand
peptides_f1<-suppressWarnings(translate(dna_seq_forward,
if.fuzzy.codon="solve"))
peptides_f2<-suppressWarnings(translate(subseq(dna_seq_forward,start=2),
if.fuzzy.codon="solve"))
peptides_f3<-suppressWarnings(translate(subseq(dna_seq_forward,start=3),
if.fuzzy.codon="solve"))
novel_cds_f1 <- .get_30aa_splited_seq(peptides_f1)
novel_cds_f2 <- .get_30aa_splited_seq(peptides_f2)
novel_cds_f3 <- .get_30aa_splited_seq(peptides_f3)
novel_cds_f1[,c("Strand","Frame","ID"):=list(a="+",
b="1",
c=paste(id,"+","F1",
Substring,
sep="|")),
by=.(id,Substring)]
novel_cds_f2[,c("Strand","Frame","ID"):=list(a="+",
b="2",
c=paste(id,"+","F2",
Substring,
sep="|")),
by=.(id,Substring)]
novel_cds_f3[,c("Strand","Frame","ID"):=list(a="+",
b="3",
c=paste(id,"+","F3",
Substring,
sep="|")),
by=.(id,Substring)]
# reverse strand
if(!bool_use_3frame)
{
dna_seq_reverse<-reverseComplement(dna_seq_forward)
peptides_r1<-suppressWarnings(translate(dna_seq_reverse,
if.fuzzy.codon="solve"))
peptides_r2<-suppressWarnings(translate(subseq(dna_seq_reverse,start=2),
if.fuzzy.codon="solve"))
peptides_r3<-suppressWarnings(translate(subseq(dna_seq_reverse,start=3),
if.fuzzy.codon="solve"))
novel_cds_r1 <- .get_30aa_splited_seq(peptides_r1)
novel_cds_r2 <- .get_30aa_splited_seq(peptides_r2)
novel_cds_r3 <- .get_30aa_splited_seq(peptides_r3)
novel_cds_r1[,c("Strand","Frame","ID"):=list(a="-",
b="1",
c=paste(id,"-","F1",
Substring,
sep="|")),
by=.(id,Substring)]
novel_cds_r2[,c("Strand","Frame","ID"):=list(a="-",
b="2",
c=paste(id,"-","F2",
Substring,
sep="|")),
by=.(id,Substring)]
novel_cds_r3[,c("Strand","Frame","ID"):=list(a="-",
b="3",
c=paste(id,"-","F3",
Substring,
sep="|")),
by=.(id,Substring)]
all_pep<-rbindlist(list(novel_cds_f1,novel_cds_f2,novel_cds_f3,
novel_cds_r1,novel_cds_r2,novel_cds_r3))
}else
{
all_pep<-rbindlist(list(novel_cds_f1,novel_cds_f2,novel_cds_f3))
}
all_pep[,Index:=paste("NTX",cumIndex+.I,sep="")]
if(bool_get_longest){
#final_all_pep<-all_pep[,.(pep=seq[nchar(seq)==max(nchar(seq))],
#ID=ID[nchar(seq)==max(nchar(seq))]),by=.(id)]
final_all_pep<-all_pep[,.(pep=seq[nchar(seq)==max(nchar(seq))],
ID=ID[nchar(seq)==max(nchar(seq))],
Index=Index[nchar(seq)==max(nchar(seq))],
Strand=Strand[nchar(seq)==max(nchar(seq))],
Frame=Frame[nchar(seq)==max(nchar(seq))],
start=start[nchar(seq)==max(nchar(seq))],
end=end[nchar(seq)==max(nchar(seq))],
Substring=Substring[nchar(seq)==max(nchar(seq))]),
by=.(id)]
if(file.exists(outmtab)){
write.table(subset(final_all_pep,
select=c(Index,id,Strand,Frame,start,end,Substring)),
file=outmtab, sep='\t', quote=FALSE, row.names=FALSE,
append=TRUE,col.names=FALSE)
}else{
write.table(subset(final_all_pep,
select=c(Index,id,Strand,Frame,start,end,Substring)),
file=outmtab, sep='\t', quote=FALSE, row.names=FALSE)
}
#final_all_pep<-all_pep[,.(pep=seq[which.max(nchar(seq))],ID=ID[which.max(nchar(seq))]),by=.(id)]
final_all_pep[,output:=paste('>',Index,"|",ID,'\n',pep,sep=''),
by=.(ID)]
write(final_all_pep$output,file=outfa,append=TRUE)
}else{
if(file.exists(outmtab)){
write.table(subset(all_pep,
select=c(Index,id,Strand,Frame,start,end,Substring)),
file=outmtab, sep='\t', quote=FALSE, row.names=FALSE,
append=TRUE,col.names=FALSE)
}else{
write.table(subset(all_pep,
select=c(Index,id,Strand,Frame,start,end,Substring)),
file=outmtab, sep='\t', quote=FALSE, row.names=FALSE)
}
all_pep[,output:=paste('>',Index,"|",ID,'\n',seq,sep=''),by=.(ID)]
write(all_pep$output,file=outfa,append=TRUE)
}
#add var tag, add decoy sequence
xset_for<-readAAStringSet(outfa)
xset_rev<-xset_for
names(xset_for)=paste(var_tag,"|",names(xset_for),sep="")
names(xset_rev)=paste(decoy_tag,var_tag,"|",names(xset_rev),sep="")
xset_rev<-reverse(xset_rev)
outfile<-paste(outfile_name,"_txFinder.fasta",sep="")
writeXStringSet(xset_for,outfile)
if(make_decoy){
writeXStringSet(xset_rev,outfile,append=TRUE)
}
message("Write protein sequences to file: ",outfile)
return(outfile)
}
.get_30aa_splited_seq<-function(translated_seq){
df<-as.data.frame(translated_seq)
df["id"]<-rownames(df)
colnames(df)<-c("raw","id")
dt<-setDT(df)
dt_split<-dt[,.(seq=unlist(strsplit(raw, "\\*"))),by=.(id,raw)]
#must be 1L, not 1
dt_split[, cumlen := cumsum((nchar(seq)+1L)), by=list(id, raw)]
dt_split[, c("start","end"):=list(cumlen-nchar(seq),cumlen-1L),
by=list(id, raw)]
#remove the pep less than 30AA .
sub.dt_split<-subset(dt_split,(end-start)>=29)
#add column "Substring"
sub.dt_split[,Substring:=seq(1,.N),by=.(id,raw)]
sub.dt_split
}
#' @title Merge the databases from RNA-Seq and the canonical reference database
#' @description Merge the databases from RNA-Seq and the canonical reference
#' database
#' @param fa A list object containing the databases from RNA-Seq
#' @param proteinseq An object containing the canonical reference database
#' @param make_decoy A logical indicating whether to add the decoy sequences
#' @param var_tag A string will be as the prefix of sequences from RNA-Seq
#' @param decoy_tag The prefix of decoy sequences
#' @param outfile_path Output directory
#' @param outfile_name Output file name
#' @return The file name of the customized database
#' @noRd
dbcat<-function(fa,proteinseq,make_decoy=TRUE,var_tag="VAR",decoy_tag="#REV#",
outfile_path=".",outfile_name="test"){
if(is.null(fa))
{
stop("Error: fasta list not found!")
}
if(is.null(proteinseq))
{
stop("Error: \"proteinseq\" not found!")
}
## process the reference proteins
## get rid of the possible "*" (stop codon) in the end of the protein
canonical_for<-AAStringSet(x=gsub("\\*$","",proteinseq$peptide))
names(canonical_for)<-proteinseq$pro_name
canonical_rev<-canonical_for
names(canonical_rev)=paste(decoy_tag,names(canonical_for),sep="")
canonical_rev<-reverse(canonical_rev)
## process the sequences from RNA-Seq
tmp_for<-tempfile(pattern="Forward_",fileext=".fasta")
tmp_rev<-tempfile(pattern="Reverse_",fileext=".fasta")
for (i in fa)
{
xset_for<-readAAStringSet(i)
xset_rev<-xset_for
names(xset_for)=paste(var_tag,"|",names(xset_for),sep="")
names(xset_rev)=paste(decoy_tag,var_tag,"|",names(xset_rev),sep="")
xset_rev<-reverse(xset_rev)
writeXStringSet(xset_for,tmp_for,append=TRUE)
writeXStringSet(xset_rev,tmp_rev,append=TRUE)
}
xset_tmp_for<-readAAStringSet(tmp_for)
xset_tmp_rev<-readAAStringSet(tmp_rev)
outfile<-paste(outfile_path,"/",outfile_name,"_txFinder.fasta",sep="")
writeXStringSet(xset_tmp_for,outfile)
writeXStringSet(canonical_for,outfile,append=TRUE)
if(make_decoy){
writeXStringSet(xset_tmp_rev,outfile,append=TRUE)
writeXStringSet(canonical_rev,outfile,append=TRUE)
}
return(outfile)
}
#' @title Add gene nemes
#' @description Add "gene name" column to the attachments (eg. snv.tsv and idl.tsv) of report
#' if there is a demand (optional function). This function is useful for "Ensembl" IDs.
#' @param mart See detail in function \code{\link{PrepareAnnotationEnsembl}}.
#' @param report The report directory for adding gene names.
#' @return none
#' @export
#' @examples
#' \dontrun{
#' library(PGA)
#' # set the biomaRt parameters just as "PrepareAnnotationEnsembl2" did
#'
#' mart <- biomaRt::useMart("ENSEMBL_MART_ENSEMBL",
#' dataset="hsapiens_gene_ensembl",
#' host="grch37.ensembl.org",
#' path="/biomart/martservice",
#' archive=FALSE)
#' addGeneName4Ensembl(mart=mart,report="report")
#'
#' }
addGeneName4Ensembl <- function(mart, report="report"){
attributes.id <- c("ensembl_peptide_id","external_gene_id")
if(file.exists(paste(report,"files/snv.tsv",sep="/"))){
file<-paste(report,"files/snv.tsv",sep="/")
raw<-read.delim(file,header=TRUE)
protein_ids<-as.character(raw$proname)
if(length(grep("^ENS",protein_ids[1]))>0)
{
idstab <- getBM(attributes=attributes.id, mart=mart,
filters='ensembl_peptide_id', values=protein_ids)
message("........ getting HGNC gene names of SNV table\n", appendLF=FALSE)
colnames(idstab)<-c("proname","geneNameHGNC")
tb <- merge(raw, idstab, by='proname',all.x=TRUE)[,union(names(raw),names(idstab))]
write.table(tb, file=paste(report,"files/snv.tsv",sep="/"),
sep='\t', quote=FALSE,row.names=FALSE)
}else
{
message("........ No conversion is required\n", appendLF=FALSE);
}
}
if(file.exists(paste(report,"files/idl.tsv",sep="/"))){
file<-paste(report,"files/idl.tsv",sep="/")
raw<-read.delim(file,header=TRUE)
protein_ids<-as.character(raw$proname)
if(length(grep("^ENS",protein_ids[1]))>0)
{
idstab <- getBM(attributes=attributes.id, mart=mart,
filters='ensembl_peptide_id', values=protein_ids)
message("........ getting HGNC gene names of InDel table\n", appendLF=FALSE)
colnames(idstab)<-c("proname","geneNameHGNC")
tb <- merge(raw, idstab, by='proname',all.x=TRUE)[,union(names(raw),names(idstab))]
write.table(tb, file=paste(report,"files/idl.tsv",sep="/"),
sep='\t', quote=FALSE,row.names=FALSE)
}else
{
message("........ No conversion is required\n", appendLF=FALSE)
}
}
}
### See https://genome.ucsc.edu/goldenpath/help/mysql.html for how to connect
### to a MySQL server at UCSC. By default UCSC_dbselect() uses the server
### located on the US west coast.
.UCSC_dbselect <- function(dbname, from, columns=NULL, where=NULL,
server="genome-mysql.soe.ucsc.edu")
{
columns <- if (is.null(columns)) "*" else paste0(columns, collapse=",")
SQL <- sprintf("SELECT %s FROM %s", columns, from)
if (!is.null(where)) {
stopifnot(isSingleString(where))
SQL <- paste(SQL, "WHERE", where)
}
dbconn <- dbConnect(RMariaDB::MariaDB(), dbname=dbname,
username="genome",
host=server,
port=3306)
on.exit(dbDisconnect(dbconn))
dbGetQuery(dbconn, SQL)
}
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