R/cnvDiscordantReads.R
In robinjugas/CNproScan: CNproScan detects and annotates CNVs in bacterial genomes.
Defines functions
cnvDiscordantReads
#' cnvDiscordantReads v2
#' Goes over BAM file and search for discordant reads. Returns input dataframe with added information.
#'
#' @importFrom Rsamtools scanBamFlag ScanBamParam scanBam
#' @importFrom GenomicRanges GRanges
#' @importFrom IRanges IRanges
#' @param bamFile path to the BAM file, sorted, indexed
#' @param CNV_DF dataframe of already detected CNVs
#' @param refHeader name of chromosome or conrig
#' @param referenceLength lengh of given contig/chromosome
#' @return CNV dataframe
#' @export
#' @noRd
#'
cnvDiscordantReads <- function(bamFile, CNV_DF, refHeader, referenceLength){
# DISCORDANT READS
## GET BASIC BAM INFO
flag<-Rsamtools::scanBamFlag(isPaired = TRUE,isUnmappedQuery=FALSE,isProperPair=TRUE)
parameters <- Rsamtools::ScanBamParam(what=c("pos", "qwidth","isize","rname"),flag=flag)
bam <- Rsamtools::scanBam(bamFile, param=parameters)[[1]]
dfBAM<-as.data.frame(bam) #create data.frame
##GET ONLY SELECTED CONTIG
dfBAM <- dfBAM[dfBAM$rname== refHeader,]
## DELETING crossovers at the start and at the end
cutoff <- 100 #defined above too
dfBAM <- dfBAM[dfBAM$pos>cutoff,]
dfBAM <- dfBAM[dfBAM$pos<(referenceLength-cutoff),]
## INSERT SIZE/TLEN - isize
insertSize <- median(abs(dfBAM$isize))
insertSize_1Q <- summary(abs(dfBAM$isize))[[2]]
insertSize_3Q <- summary(abs(dfBAM$isize))[[5]]
insertSize_IQR <- IQR(abs(dfBAM$isize))
## READ LENGTH - qwidth: The width of the query, as calculated from the cigar encoding; normally equal to the width of the query returned in seq.
readLength <- median(abs(dfBAM$qwidth))
################################################################################
## SCAN BAM in CNV REGIONS
if(nrow(CNV_DF)>0){
for(i in 1:nrow(CNV_DF)){
start <- CNV_DF[i,"START"]
stop <- CNV_DF[i,"END"]
# Granges boundaries
STARTGRANGES <- start-insertSize
STOPGRANGES <- stop+insertSize
if (STARTGRANGES<1){STARTGRANGES <- 1}
if(STOPGRANGES>referenceLength){STOPGRANGES <- referenceLength}
region <- GenomicRanges::GRanges(refHeader, IRanges::IRanges(start-insertSize, stop+insertSize))
CNVLENGTH <- CNV_DF[i,"LENGTH"]
#-----------------------------------------------------------------------------
## ACCESS BAM - TOTAL READS COUNT
flagTOTAL <- Rsamtools::scanBamFlag(isPaired = TRUE)
parametersTOTAL <- Rsamtools::ScanBamParam(what=c("pos", "qwidth","isize"),which=region, flag=flagTOTAL)
bamTOTAL <- Rsamtools::scanBam(bamFile, param=parametersTOTAL)[[1]]
dfBAMTOTAL <- as.data.frame(bamTOTAL)
dfBAMTOTAL <- na.omit(dfBAMTOTAL)
if(!is.null(nrow(dfBAMTOTAL))){reads_TOTAL <- nrow(dfBAMTOTAL)}else{reads_TOTAL <- 0}
################################################################################################################################################################
#-----------------------------------------------------------------------------
## ACCESS BAM - DELETIONS - reads orientation +/- -/+ AND higher INSERT SIZE
# reads orientation +/-
flagDel1 <- Rsamtools::scanBamFlag(isPaired = TRUE, isMinusStrand=FALSE, isFirstMateRead=TRUE,isMateMinusStrand=TRUE) # reads orientation +/-
parametersDel1 <- Rsamtools::ScanBamParam(what=c("pos", "qwidth","isize"),which=region, flag=flagDel1)
dfBAM <- Rsamtools::scanBam(bamFile, param=parametersDel1)[[1]]
dfBAM <- as.data.frame(dfBAM)
dfBAM <- na.omit(dfBAM)
# %% Circular genome correction
if(!is.null(nrow(dfBAM))){
# % positive values
REF <- referenceLength/2
dfBAM[dfBAM$isize>REF,"isize"] <- dfBAM[dfBAM$isize>REF,"isize"] - 2*dfBAM[dfBAM$isize>REF,"qwidth"] - referenceLength
# % negative values
REF <- referenceLength/-2
dfBAM[dfBAM$isize<REF,"isize"] <- dfBAM[dfBAM$isize<REF,"isize"] + 2*dfBAM[dfBAM$isize<REF,"qwidth"] + referenceLength
#find outliers in TLEN fragment size 1.5xIQR rule or as same as CNV length
out_ind <- which(abs(dfBAM$isize)>(insertSize_3Q+1.5*insertSize_IQR))
dfBAM_INCREASE<-dfBAM[out_ind,]
out_ind <- which(abs(dfBAM$isize)>(CNVLENGTH-2*insertSize) & abs(dfBAM$isize)<(CNVLENGTH+2*insertSize))
dfBAM_ASCNVLENGTH<-dfBAM[out_ind,]
}
if(!is.null(nrow(dfBAM_INCREASE))){reads_supporting_DELETION1 <- nrow(dfBAM_INCREASE)}else{reads_supporting_DELETION1 <- 0}
if(!is.null(nrow(dfBAM_ASCNVLENGTH)) & !is.null(nrow(dfBAM_INCREASE))){reads_supporting_DELETION1 <- reads_supporting_DELETION1+nrow(dfBAM_ASCNVLENGTH)}
# reads orientation -/+
flagDel2 <- Rsamtools::scanBamFlag(isPaired = TRUE, isMinusStrand=TRUE, isFirstMateRead=TRUE,isMateMinusStrand=FALSE)
parametersDel2 <- Rsamtools::ScanBamParam(what=c("pos", "qwidth","isize"),which=region, flag=flagDel2)
dfBAM <- Rsamtools::scanBam(bamFile, param=parametersDel2)[[1]]
dfBAM <- as.data.frame(dfBAM)
dfBAM <- na.omit(dfBAM)
# %% Circular genome correction
if(!is.null(nrow(dfBAM))){
# % positive values
REF <- referenceLength/2
dfBAM[dfBAM$isize>REF,"isize"] <- dfBAM[dfBAM$isize>REF,"isize"] - 2*dfBAM[dfBAM$isize>REF,"qwidth"] - referenceLength
# % negative values
REF <- referenceLength/-2
dfBAM[dfBAM$isize<REF,"isize"] <- dfBAM[dfBAM$isize<REF,"isize"] + 2*dfBAM[dfBAM$isize<REF,"qwidth"] + referenceLength
#find outliers in TLEN fragment size 1.5xIQR rule or as same as CNV length
out_ind <- which(abs(dfBAM$isize)>(insertSize_3Q+1.5*insertSize_IQR))
dfBAM_INCREASE<-dfBAM[out_ind,]
out_ind <- which(abs(dfBAM$isize)>(CNVLENGTH-2*insertSize) & abs(dfBAM$isize)<(CNVLENGTH+2*insertSize))
dfBAM_ASCNVLENGTH<-dfBAM[out_ind,]
}
if(!is.null(nrow(dfBAM_INCREASE))){reads_supporting_DELETION2 <- nrow(dfBAM_INCREASE)}else{reads_supporting_DELETION2 <- 0}
if(!is.null(nrow(dfBAM_ASCNVLENGTH)) & !is.null(nrow(dfBAM_INCREASE))){reads_supporting_DELETION2 <- reads_supporting_DELETION2+nrow(dfBAM_ASCNVLENGTH)}
reads_supporting_DELETION <- reads_supporting_DELETION1 + reads_supporting_DELETION2
################################################################################################################################################################
#-----------------------------------------------------------------------------
## ACCESS BAM TANDEM DUPLICATIONS DIRECT - reads orientation -/+ or +/- AND lower INSERT SIZE OR INCREASE AROUND SIZE of CNV length +- insertSize
# reads orientation -/+
flagTD_DIR1 <- Rsamtools::scanBamFlag(isPaired = TRUE, isMinusStrand=TRUE, isFirstMateRead=TRUE,isMateMinusStrand=FALSE)
parametersTD_DIR1 <- Rsamtools::ScanBamParam(what = c("pos", "qwidth","isize"), which=region, flag=flagTD_DIR1)
dfBAM <- Rsamtools::scanBam(bamFile, param=parametersTD_DIR1)[[1]]
dfBAM <- as.data.frame(dfBAM)
dfBAM <- na.omit(dfBAM)
# %% Circular genome correction
if(!is.null(nrow(dfBAM))){
# % positive values
REF <- referenceLength/2
dfBAM[dfBAM$isize>REF,"isize"] <- dfBAM[dfBAM$isize>REF,"isize"] - 2*dfBAM[dfBAM$isize>REF,"qwidth"] - referenceLength
# % negative values
REF <- referenceLength/-2
dfBAM[dfBAM$isize<REF,"isize"] <- dfBAM[dfBAM$isize<REF,"isize"] + 2*dfBAM[dfBAM$isize<REF,"qwidth"] + referenceLength
#find outliers in TLEN fragment size 1.5xIQR rule or as same as CNV length
out_ind <- which(abs(dfBAM$isize)>(CNVLENGTH-2*insertSize) & abs(dfBAM$isize)<(CNVLENGTH+2*insertSize))
dfBAM_ASCNVLENGTH<-dfBAM[out_ind,]
out_ind <- which(abs(dfBAM$isize)<(insertSize_1Q-1.5*insertSize_IQR))
dfBAM_DECREASE<-dfBAM[out_ind,]
}
if(!is.null(nrow(dfBAM_DECREASE))){reads_supporting_TADDIR1 <- nrow(dfBAM_DECREASE)}else{reads_supporting_TADDIR1 <- 0}
if(!is.null(nrow(dfBAM_ASCNVLENGTH)) & !is.null(nrow(dfBAM_DECREASE))){reads_supporting_TADDIR1 <- reads_supporting_TADDIR1+nrow(dfBAM_ASCNVLENGTH)}
# reads orientation +/-
flagTD_DIR2 <- Rsamtools::scanBamFlag(isPaired = TRUE, isMinusStrand=TRUE, isFirstMateRead=TRUE,isMateMinusStrand=FALSE)
parametersTD_DIR2 <- Rsamtools::ScanBamParam(what = c("pos", "qwidth","isize"), which=region, flag=flagTD_DIR2)
dfBAM <- Rsamtools::scanBam(bamFile, param=parametersTD_DIR2)[[1]]
dfBAM <- as.data.frame(dfBAM)
dfBAM <- na.omit(dfBAM)
# %% Circular genome correction
if(!is.null(nrow(dfBAM))){
# % positive values
REF <- referenceLength/2
dfBAM[dfBAM$isize>REF,"isize"] <- dfBAM[dfBAM$isize>REF,"isize"] - 2*dfBAM[dfBAM$isize>REF,"qwidth"] - referenceLength
# % negative values
REF <- referenceLength/-2
dfBAM[dfBAM$isize<REF,"isize"] <- dfBAM[dfBAM$isize<REF,"isize"] + 2*dfBAM[dfBAM$isize<REF,"qwidth"] + referenceLength
#find outliers in TLEN fragment size 1.5xIQR rule or as same as CNV length
out_ind <- which(abs(dfBAM$isize)>(CNVLENGTH-2*insertSize) & abs(dfBAM$isize)<(CNVLENGTH+2*insertSize))
dfBAM_ASCNVLENGTH<-dfBAM[out_ind,]
out_ind <- which(abs(dfBAM$isize)<(insertSize_1Q-1.5*insertSize_IQR))
dfBAM_DECREASE<-dfBAM[out_ind,]
}
if(!is.null(nrow(dfBAM_DECREASE))){reads_supporting_TADDIR2 <- nrow(dfBAM_DECREASE)}else{reads_supporting_TADDIR2 <- 0}
if(!is.null(nrow(dfBAM_ASCNVLENGTH)) & !is.null(nrow(dfBAM_DECREASE))){reads_supporting_TADDIR2 <- reads_supporting_TADDIR2+nrow(dfBAM_ASCNVLENGTH)}
reads_supporting_TANDEM_DIRECT <- reads_supporting_TADDIR1 + reads_supporting_TADDIR2
#-----------------------------------------------------------------------------
## ACCESS BAM TANDEM DUPLICATIONS INDIRECT - reads orientation -/- or +/+ AND lower INSERT SIZE
# reads orientation -/-
flagTD_INDIR1 <- Rsamtools::scanBamFlag(isPaired = TRUE, isMinusStrand=TRUE, isMateMinusStrand=TRUE)
parametersTD_INDIR1 <- Rsamtools::ScanBamParam(what = c("pos", "qwidth","isize"), which=region, flag=flagTD_INDIR1)
dfBAM <- Rsamtools::scanBam(bamFile, param=parametersTD_INDIR1)[[1]]
dfBAM <- as.data.frame(dfBAM)
dfBAM <- na.omit(dfBAM)
# %% Circular genome correction
if(!is.null(nrow(dfBAM))){
# % positive values
REF <- referenceLength/2
dfBAM[dfBAM$isize>REF,"isize"] <- dfBAM[dfBAM$isize>REF,"isize"] - 2*dfBAM[dfBAM$isize>REF,"qwidth"] - referenceLength
# % negative values
REF <- referenceLength/-2
dfBAM[dfBAM$isize<REF,"isize"] <- dfBAM[dfBAM$isize<REF,"isize"] + 2*dfBAM[dfBAM$isize<REF,"qwidth"] + referenceLength
#find outliers in TLEN fragment size 1.5xIQR rule or as same as CNV length
out_ind <- which(abs(dfBAM$isize)>(CNVLENGTH-2*insertSize) & abs(dfBAM$isize)<(CNVLENGTH+2*insertSize))
dfBAM_ASCNVLENGTH<-dfBAM[out_ind,]
out_ind <- which(abs(dfBAM$isize)<(insertSize_1Q-1.5*insertSize_IQR))
dfBAM_DECREASE<-dfBAM[out_ind,]
}
if(!is.null(nrow(dfBAM_DECREASE))){reads_supporting_TADINDIR1 <- nrow(dfBAM_DECREASE)}else{reads_supporting_TADINDIR1 <- 0}
if(!is.null(nrow(dfBAM_ASCNVLENGTH)) & !is.null(nrow(dfBAM_DECREASE))){reads_supporting_TADINDIR1 <- reads_supporting_TADINDIR1+nrow(dfBAM_ASCNVLENGTH)}
# reads orientation +/+
flagTD_INDIR2 <- Rsamtools::scanBamFlag(isPaired = TRUE, isMinusStrand=FALSE, isMateMinusStrand=FALSE)
parametersTD_INDIR2 <- Rsamtools::ScanBamParam(what = c("pos", "qwidth","isize"), which=region, flag=flagTD_INDIR2)
dfBAM <- Rsamtools::scanBam(bamFile, param=parametersTD_INDIR2)[[1]]
dfBAM <- as.data.frame(dfBAM)
dfBAM <- na.omit(dfBAM)
# %% Circular genome correction
if(!is.null(nrow(dfBAM))){
# % positive values
REF <- referenceLength/2
dfBAM[dfBAM$isize>REF,"isize"] <- dfBAM[dfBAM$isize>REF,"isize"] - 2*dfBAM[dfBAM$isize>REF,"qwidth"] - referenceLength
# % negative values
REF <- referenceLength/-2
dfBAM[dfBAM$isize<REF,"isize"] <- dfBAM[dfBAM$isize<REF,"isize"] + 2*dfBAM[dfBAM$isize<REF,"qwidth"] + referenceLength
#find outliers in TLEN fragment size 1.5xIQR rule or as same as CNV length
out_ind <- which(abs(dfBAM$isize)>(CNVLENGTH-2*insertSize) & abs(dfBAM$isize)<(CNVLENGTH+2*insertSize))
dfBAM_ASCNVLENGTH<-dfBAM[out_ind,]
out_ind <- which(abs(dfBAM$isize)<(insertSize_1Q-1.5*insertSize_IQR))
dfBAM_DECREASE<-dfBAM[out_ind,]
}
if(!is.null(nrow(dfBAM_DECREASE))){reads_supporting_TADINDIR2 <- nrow(dfBAM_DECREASE)}else{reads_supporting_TADINDIR2 <- 0}
if(!is.null(nrow(dfBAM_ASCNVLENGTH)) & !is.null(nrow(dfBAM_DECREASE))){reads_supporting_TADINDIR2 <- reads_supporting_TADINDIR2+nrow(dfBAM_ASCNVLENGTH)}
reads_supporting_TANDEM_INDIRECT <- reads_supporting_TADINDIR1 + reads_supporting_TADINDIR2
################################################################################################################################################################
#-----------------------------------------------------------------------------
# ACCESS BAM INTERSPERSED DUPLICATIONS INVERSED - reads orientation -/- OR +/+ AND much higher INSERT SIZE
# reads orientation -/-
flagINTERDUP_INV1 <- Rsamtools::scanBamFlag(isPaired = TRUE, isMinusStrand=TRUE, isMateMinusStrand=TRUE)
parametersINTERDUP_INV1 <- Rsamtools::ScanBamParam(what = c("pos", "qwidth","isize"), which=region, flag=flagINTERDUP_INV1)
dfBAM <- Rsamtools::scanBam(bamFile, param=parametersINTERDUP_INV1)[[1]]
dfBAM <- as.data.frame(dfBAM)
dfBAM <- na.omit(dfBAM)
# %% Circular genome correction
if(!is.null(nrow(dfBAM))){
# % positive values
REF <- referenceLength/2
dfBAM[dfBAM$isize>REF,"isize"] <- dfBAM[dfBAM$isize>REF,"isize"] - 2*dfBAM[dfBAM$isize>REF,"qwidth"] - referenceLength
# % negative values
REF <- referenceLength/-2
dfBAM[dfBAM$isize<REF,"isize"] <- dfBAM[dfBAM$isize<REF,"isize"] + 2*dfBAM[dfBAM$isize<REF,"qwidth"] + referenceLength
#find outliers in TLEN fragment size 1.5xIQR rule
out_ind <- which(abs(dfBAM$isize)>(insertSize_3Q+10*insertSize_IQR))
dfBAM_INCREASE<-dfBAM[out_ind,]
}
if(!is.null(nrow(dfBAM_INCREASE))){reads_count_INTERDUP_INV1 <- nrow(dfBAM_INCREASE)}else{reads_count_INTERDUP_INV1 <- 0}
# reads orientation +/+
flagINTERDUP_INV2 <- Rsamtools::scanBamFlag(isPaired = TRUE, isMinusStrand=FALSE, isMateMinusStrand=FALSE)
parametersINTERDUP_INV2 <- Rsamtools::ScanBamParam(what = c("pos", "qwidth","isize"),which=region,flag=flagINTERDUP_INV2)
dfBAM <- Rsamtools::scanBam(bamFile, param=parametersINTERDUP_INV2)[[1]]
dfBAM <- as.data.frame(dfBAM)
dfBAM <- na.omit(dfBAM)
# %% Circular genome correction
if(!is.null(nrow(dfBAM))){
# % positive values
REF <- referenceLength/2
dfBAM[dfBAM$isize>REF,"isize"] <- dfBAM[dfBAM$isize>REF,"isize"] - 2*dfBAM[dfBAM$isize>REF,"qwidth"] - referenceLength
# % negative values
REF <- referenceLength/-2
dfBAM[dfBAM$isize<REF,"isize"] <- dfBAM[dfBAM$isize<REF,"isize"] + 2*dfBAM[dfBAM$isize<REF,"qwidth"] + referenceLength
#find outliers in TLEN fragment size 1.5xIQR rule
out_ind <- which(abs(dfBAM$isize)>(insertSize_3Q+10*insertSize_IQR))
dfBAM_INCREASE<-dfBAM[out_ind,]
}
if(!is.null(nrow(dfBAM_INCREASE))){reads_count_INTERDUP_INV2 <- nrow(dfBAM_INCREASE)}else{reads_count_INTERDUP_INV2 <- 0}
reads_supporting_INTERSPERSED_INDIRECT <- reads_count_INTERDUP_INV2 + reads_count_INTERDUP_INV1
#-----------------------------------------------------------------------------
# ACCESS BAM INTERSPERSED DUPLICATIONS DIRECT - reads orientation +/- OR -/+ AND much higher INSERT SIZE
# reads orientation +/-
flagINTERDUP_DIR1 <- Rsamtools::scanBamFlag(isPaired = TRUE, isMinusStrand=FALSE, isFirstMateRead=TRUE,isMateMinusStrand=TRUE)
parametersINTERDUP_DIR1 <- Rsamtools::ScanBamParam(what = c("pos", "qwidth","isize"), which=region, flag=flagINTERDUP_DIR1)
dfBAM <- Rsamtools::scanBam(bamFile, param=parametersINTERDUP_DIR1)[[1]]
dfBAM <- as.data.frame(dfBAM)
dfBAM <- na.omit(dfBAM)
# %% Circular genome correction
if(!is.null(nrow(dfBAM))){
# % positive values
REF <- referenceLength/2
dfBAM[dfBAM$isize>REF,"isize"] <- dfBAM[dfBAM$isize>REF,"isize"] - 2*dfBAM[dfBAM$isize>REF,"qwidth"] - referenceLength
# % negative values
REF <- referenceLength/-2
dfBAM[dfBAM$isize<REF,"isize"] <- dfBAM[dfBAM$isize<REF,"isize"] + 2*dfBAM[dfBAM$isize<REF,"qwidth"] + referenceLength
#find outliers in TLEN fragment size 1.5xIQR rule
out_ind <- which(abs(dfBAM$isize)>(insertSize_3Q+10*insertSize_IQR))
dfBAM_INCREASE<-dfBAM[out_ind,]
}
if(!is.null(nrow(dfBAM_INCREASE))){reads_count_INTERDUP_DIR1 <- nrow(dfBAM_INCREASE)}else{reads_count_INTERDUP_DIR1 <- 0}
# reads orientation -/+
flagINTERDUP_DIR2 <- Rsamtools::scanBamFlag(isPaired = TRUE, isMinusStrand=TRUE, isFirstMateRead=TRUE,isMateMinusStrand=FALSE)
parametersINTERDUP_DIR2 <- Rsamtools::ScanBamParam(what = c("pos", "qwidth","isize"),which=region,flag=flagINTERDUP_DIR2)
dfBAM <- Rsamtools::scanBam(bamFile, param=parametersINTERDUP_DIR2)[[1]]
dfBAM <- as.data.frame(dfBAM)
dfBAM <- na.omit(dfBAM)
# %% Circular genome correction
if(!is.null(nrow(dfBAM))){
# % positive values
REF <- referenceLength/2
dfBAM[dfBAM$isize>REF,"isize"] <- dfBAM[dfBAM$isize>REF,"isize"] - 2*dfBAM[dfBAM$isize>REF,"qwidth"] - referenceLength
# % negative values
REF <- referenceLength/-2
dfBAM[dfBAM$isize<REF,"isize"] <- dfBAM[dfBAM$isize<REF,"isize"] + 2*dfBAM[dfBAM$isize<REF,"qwidth"] + referenceLength
#find outliers in TLEN fragment size 1.5xIQR rule
out_ind <- which(abs(dfBAM$isize)>(insertSize_3Q+10*insertSize_IQR))
dfBAM_INCREASE<-dfBAM[out_ind,]
}
if(!is.null(nrow(dfBAM_INCREASE))){reads_count_INTERDUP_DIR2 <- nrow(dfBAM_INCREASE)}else{reads_count_INTERDUP_DIR2 <- 0}
reads_supporting_INTERSPERSED_DIRECT <- reads_count_INTERDUP_DIR2 + reads_count_INTERDUP_DIR1
#-----------------------------------------------------------------------------
# #print absolute numbers
CNV_DF[i,"reads_TOTAL"] <- reads_TOTAL
CNV_DF[i,"reads_supporting_Deletion"] <- reads_supporting_DELETION
CNV_DF[i,"reads_supporting_Tandem_Direct"] <- reads_supporting_TANDEM_DIRECT
CNV_DF[i,"reads_supporting_Tandem_Indirect"] <- reads_supporting_TANDEM_INDIRECT
CNV_DF[i,"reads_supporting_Interspersed_Direct"] <- reads_supporting_INTERSPERSED_DIRECT
CNV_DF[i,"reads_supporting_Interspersed_Indirect"] <- reads_supporting_INTERSPERSED_INDIRECT
#print Percentages
# CNV_DF[i,"reads_TOTAL"] <- reads_TOTAL
# CNV_DF[i,"reads_INSERT_INCREASE"] <- reads_count_INCREASE/(reads_TOTAL/100)
# CNV_DF[i,"reads_INSERT_DECREASE"] <- reads_count_DECREASE/(reads_TOTAL/100)
# CNV_DF[i,"reads_count_TAD_DUP"] <- reads_count_TD/(reads_TOTAL/100)
# CNV_DF[i,"reads_count_INTER_DUP_DIR"] <- reads_count_INTERDUP_DIR/(reads_TOTAL/100)
# CNV_DF[i,"reads_count_INTER_DUP_INV"] <- reads_count_INTERDUP_INV/(reads_TOTAL/100)
# SUBTYPE SELECTION
signaturesSUM <- reads_supporting_DELETION+reads_supporting_TANDEM_DIRECT+reads_supporting_TANDEM_INDIRECT+reads_supporting_INTERSPERSED_DIRECT+reads_supporting_INTERSPERSED_INDIRECT
tempX <- which.max(c(reads_supporting_TANDEM_DIRECT,reads_supporting_TANDEM_INDIRECT,reads_supporting_INTERSPERSED_DIRECT,reads_supporting_INTERSPERSED_INDIRECT))
if(CNV_DF[i,"TYPE"] == "DEL") {CNV_DF[i,"SUBTYPE"] <- "DEL"}
if(CNV_DF[i,"TYPE"] == "DUP" & tempX==1) {CNV_DF[i,"SUBTYPE"] <- "TANDEM_DIRECT"}
if(CNV_DF[i,"TYPE"] == "DUP" & tempX==2) {CNV_DF[i,"SUBTYPE"] <- "TANDEM_INDIRECT"}
if(CNV_DF[i,"TYPE"] == "DUP" & tempX==3) {CNV_DF[i,"SUBTYPE"] <- "INTERSPERSED_DIRECT"}
if(CNV_DF[i,"TYPE"] == "DUP" & tempX==4) {CNV_DF[i,"SUBTYPE"] <- "INTERSPERSED_INDIRECT"}
if(signaturesSUM < (reads_TOTAL/10) ) {CNV_DF[i,"SUBTYPE"] <- "FALSE POSITIVE"}
#reorder
CNV_DF <- CNV_DF[,c("ID",
"START",
"END",
"LENGTH",
"COVERAGE",
"TYPE",
"SUBTYPE",
"reads_TOTAL",
"reads_supporting_Deletion",
"reads_supporting_Tandem_Direct",
"reads_supporting_Tandem_Indirect",
"reads_supporting_Interspersed_Direct",
"reads_supporting_Interspersed_Indirect"
)]
}
} else{CNV_DF <- data.frame(ID=as.character(), START=integer(), END=integer(), LENGTH=integer(), COVERAGE=integer(), TYPE=character(),SUBTYPE=as.character(),
reads_TOTAL=integer(),reads_supporting_Deletion=integer(),reads_supporting_Tandem_Direct=integer(),reads_supporting_Tandem_Indirect=integer(),
reads_supporting_Interspersed_Direct=integer(),reads_supporting_Interspersed_Indirect=integer())
}
return(CNV_DF)
}
robinjugas/CNproScan documentation built on Jan. 11, 2025, 10:36 a.m.
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Defines functions cnvDiscordantReads
#' cnvDiscordantReads v2
#' Goes over BAM file and search for discordant reads. Returns input dataframe with added information.
#'
#' @importFrom Rsamtools scanBamFlag ScanBamParam scanBam
#' @importFrom GenomicRanges GRanges
#' @importFrom IRanges IRanges
#' @param bamFile path to the BAM file, sorted, indexed
#' @param CNV_DF dataframe of already detected CNVs
#' @param refHeader name of chromosome or conrig
#' @param referenceLength lengh of given contig/chromosome
#' @return CNV dataframe
#' @export
#' @noRd
#'
cnvDiscordantReads <- function(bamFile, CNV_DF, refHeader, referenceLength){
# DISCORDANT READS
## GET BASIC BAM INFO
flag<-Rsamtools::scanBamFlag(isPaired = TRUE,isUnmappedQuery=FALSE,isProperPair=TRUE)
parameters <- Rsamtools::ScanBamParam(what=c("pos", "qwidth","isize","rname"),flag=flag)
bam <- Rsamtools::scanBam(bamFile, param=parameters)[[1]]
dfBAM<-as.data.frame(bam) #create data.frame
##GET ONLY SELECTED CONTIG
dfBAM <- dfBAM[dfBAM$rname== refHeader,]
## DELETING crossovers at the start and at the end
cutoff <- 100 #defined above too
dfBAM <- dfBAM[dfBAM$pos>cutoff,]
dfBAM <- dfBAM[dfBAM$pos<(referenceLength-cutoff),]
## INSERT SIZE/TLEN - isize
insertSize <- median(abs(dfBAM$isize))
insertSize_1Q <- summary(abs(dfBAM$isize))[[2]]
insertSize_3Q <- summary(abs(dfBAM$isize))[[5]]
insertSize_IQR <- IQR(abs(dfBAM$isize))
## READ LENGTH - qwidth: The width of the query, as calculated from the cigar encoding; normally equal to the width of the query returned in seq.
readLength <- median(abs(dfBAM$qwidth))
################################################################################
## SCAN BAM in CNV REGIONS
if(nrow(CNV_DF)>0){
for(i in 1:nrow(CNV_DF)){
start <- CNV_DF[i,"START"]
stop <- CNV_DF[i,"END"]
# Granges boundaries
STARTGRANGES <- start-insertSize
STOPGRANGES <- stop+insertSize
if (STARTGRANGES<1){STARTGRANGES <- 1}
if(STOPGRANGES>referenceLength){STOPGRANGES <- referenceLength}
region <- GenomicRanges::GRanges(refHeader, IRanges::IRanges(start-insertSize, stop+insertSize))
CNVLENGTH <- CNV_DF[i,"LENGTH"]
#-----------------------------------------------------------------------------
## ACCESS BAM - TOTAL READS COUNT
flagTOTAL <- Rsamtools::scanBamFlag(isPaired = TRUE)
parametersTOTAL <- Rsamtools::ScanBamParam(what=c("pos", "qwidth","isize"),which=region, flag=flagTOTAL)
bamTOTAL <- Rsamtools::scanBam(bamFile, param=parametersTOTAL)[[1]]
dfBAMTOTAL <- as.data.frame(bamTOTAL)
dfBAMTOTAL <- na.omit(dfBAMTOTAL)
if(!is.null(nrow(dfBAMTOTAL))){reads_TOTAL <- nrow(dfBAMTOTAL)}else{reads_TOTAL <- 0}
################################################################################################################################################################
#-----------------------------------------------------------------------------
## ACCESS BAM - DELETIONS - reads orientation +/- -/+ AND higher INSERT SIZE
# reads orientation +/-
flagDel1 <- Rsamtools::scanBamFlag(isPaired = TRUE, isMinusStrand=FALSE, isFirstMateRead=TRUE,isMateMinusStrand=TRUE) # reads orientation +/-
parametersDel1 <- Rsamtools::ScanBamParam(what=c("pos", "qwidth","isize"),which=region, flag=flagDel1)
dfBAM <- Rsamtools::scanBam(bamFile, param=parametersDel1)[[1]]
dfBAM <- as.data.frame(dfBAM)
dfBAM <- na.omit(dfBAM)
# %% Circular genome correction
if(!is.null(nrow(dfBAM))){
# % positive values
REF <- referenceLength/2
dfBAM[dfBAM$isize>REF,"isize"] <- dfBAM[dfBAM$isize>REF,"isize"] - 2*dfBAM[dfBAM$isize>REF,"qwidth"] - referenceLength
# % negative values
REF <- referenceLength/-2
dfBAM[dfBAM$isize<REF,"isize"] <- dfBAM[dfBAM$isize<REF,"isize"] + 2*dfBAM[dfBAM$isize<REF,"qwidth"] + referenceLength
#find outliers in TLEN fragment size 1.5xIQR rule or as same as CNV length
out_ind <- which(abs(dfBAM$isize)>(insertSize_3Q+1.5*insertSize_IQR))
dfBAM_INCREASE<-dfBAM[out_ind,]
out_ind <- which(abs(dfBAM$isize)>(CNVLENGTH-2*insertSize) & abs(dfBAM$isize)<(CNVLENGTH+2*insertSize))
dfBAM_ASCNVLENGTH<-dfBAM[out_ind,]
}
if(!is.null(nrow(dfBAM_INCREASE))){reads_supporting_DELETION1 <- nrow(dfBAM_INCREASE)}else{reads_supporting_DELETION1 <- 0}
if(!is.null(nrow(dfBAM_ASCNVLENGTH)) & !is.null(nrow(dfBAM_INCREASE))){reads_supporting_DELETION1 <- reads_supporting_DELETION1+nrow(dfBAM_ASCNVLENGTH)}
# reads orientation -/+
flagDel2 <- Rsamtools::scanBamFlag(isPaired = TRUE, isMinusStrand=TRUE, isFirstMateRead=TRUE,isMateMinusStrand=FALSE)
parametersDel2 <- Rsamtools::ScanBamParam(what=c("pos", "qwidth","isize"),which=region, flag=flagDel2)
dfBAM <- Rsamtools::scanBam(bamFile, param=parametersDel2)[[1]]
dfBAM <- as.data.frame(dfBAM)
dfBAM <- na.omit(dfBAM)
# %% Circular genome correction
if(!is.null(nrow(dfBAM))){
# % positive values
REF <- referenceLength/2
dfBAM[dfBAM$isize>REF,"isize"] <- dfBAM[dfBAM$isize>REF,"isize"] - 2*dfBAM[dfBAM$isize>REF,"qwidth"] - referenceLength
# % negative values
REF <- referenceLength/-2
dfBAM[dfBAM$isize<REF,"isize"] <- dfBAM[dfBAM$isize<REF,"isize"] + 2*dfBAM[dfBAM$isize<REF,"qwidth"] + referenceLength
#find outliers in TLEN fragment size 1.5xIQR rule or as same as CNV length
out_ind <- which(abs(dfBAM$isize)>(insertSize_3Q+1.5*insertSize_IQR))
dfBAM_INCREASE<-dfBAM[out_ind,]
out_ind <- which(abs(dfBAM$isize)>(CNVLENGTH-2*insertSize) & abs(dfBAM$isize)<(CNVLENGTH+2*insertSize))
dfBAM_ASCNVLENGTH<-dfBAM[out_ind,]
}
if(!is.null(nrow(dfBAM_INCREASE))){reads_supporting_DELETION2 <- nrow(dfBAM_INCREASE)}else{reads_supporting_DELETION2 <- 0}
if(!is.null(nrow(dfBAM_ASCNVLENGTH)) & !is.null(nrow(dfBAM_INCREASE))){reads_supporting_DELETION2 <- reads_supporting_DELETION2+nrow(dfBAM_ASCNVLENGTH)}
reads_supporting_DELETION <- reads_supporting_DELETION1 + reads_supporting_DELETION2
################################################################################################################################################################
#-----------------------------------------------------------------------------
## ACCESS BAM TANDEM DUPLICATIONS DIRECT - reads orientation -/+ or +/- AND lower INSERT SIZE OR INCREASE AROUND SIZE of CNV length +- insertSize
# reads orientation -/+
flagTD_DIR1 <- Rsamtools::scanBamFlag(isPaired = TRUE, isMinusStrand=TRUE, isFirstMateRead=TRUE,isMateMinusStrand=FALSE)
parametersTD_DIR1 <- Rsamtools::ScanBamParam(what = c("pos", "qwidth","isize"), which=region, flag=flagTD_DIR1)
dfBAM <- Rsamtools::scanBam(bamFile, param=parametersTD_DIR1)[[1]]
dfBAM <- as.data.frame(dfBAM)
dfBAM <- na.omit(dfBAM)
# %% Circular genome correction
if(!is.null(nrow(dfBAM))){
# % positive values
REF <- referenceLength/2
dfBAM[dfBAM$isize>REF,"isize"] <- dfBAM[dfBAM$isize>REF,"isize"] - 2*dfBAM[dfBAM$isize>REF,"qwidth"] - referenceLength
# % negative values
REF <- referenceLength/-2
dfBAM[dfBAM$isize<REF,"isize"] <- dfBAM[dfBAM$isize<REF,"isize"] + 2*dfBAM[dfBAM$isize<REF,"qwidth"] + referenceLength
#find outliers in TLEN fragment size 1.5xIQR rule or as same as CNV length
out_ind <- which(abs(dfBAM$isize)>(CNVLENGTH-2*insertSize) & abs(dfBAM$isize)<(CNVLENGTH+2*insertSize))
dfBAM_ASCNVLENGTH<-dfBAM[out_ind,]
out_ind <- which(abs(dfBAM$isize)<(insertSize_1Q-1.5*insertSize_IQR))
dfBAM_DECREASE<-dfBAM[out_ind,]
}
if(!is.null(nrow(dfBAM_DECREASE))){reads_supporting_TADDIR1 <- nrow(dfBAM_DECREASE)}else{reads_supporting_TADDIR1 <- 0}
if(!is.null(nrow(dfBAM_ASCNVLENGTH)) & !is.null(nrow(dfBAM_DECREASE))){reads_supporting_TADDIR1 <- reads_supporting_TADDIR1+nrow(dfBAM_ASCNVLENGTH)}
# reads orientation +/-
flagTD_DIR2 <- Rsamtools::scanBamFlag(isPaired = TRUE, isMinusStrand=TRUE, isFirstMateRead=TRUE,isMateMinusStrand=FALSE)
parametersTD_DIR2 <- Rsamtools::ScanBamParam(what = c("pos", "qwidth","isize"), which=region, flag=flagTD_DIR2)
dfBAM <- Rsamtools::scanBam(bamFile, param=parametersTD_DIR2)[[1]]
dfBAM <- as.data.frame(dfBAM)
dfBAM <- na.omit(dfBAM)
# %% Circular genome correction
if(!is.null(nrow(dfBAM))){
# % positive values
REF <- referenceLength/2
dfBAM[dfBAM$isize>REF,"isize"] <- dfBAM[dfBAM$isize>REF,"isize"] - 2*dfBAM[dfBAM$isize>REF,"qwidth"] - referenceLength
# % negative values
REF <- referenceLength/-2
dfBAM[dfBAM$isize<REF,"isize"] <- dfBAM[dfBAM$isize<REF,"isize"] + 2*dfBAM[dfBAM$isize<REF,"qwidth"] + referenceLength
#find outliers in TLEN fragment size 1.5xIQR rule or as same as CNV length
out_ind <- which(abs(dfBAM$isize)>(CNVLENGTH-2*insertSize) & abs(dfBAM$isize)<(CNVLENGTH+2*insertSize))
dfBAM_ASCNVLENGTH<-dfBAM[out_ind,]
out_ind <- which(abs(dfBAM$isize)<(insertSize_1Q-1.5*insertSize_IQR))
dfBAM_DECREASE<-dfBAM[out_ind,]
}
if(!is.null(nrow(dfBAM_DECREASE))){reads_supporting_TADDIR2 <- nrow(dfBAM_DECREASE)}else{reads_supporting_TADDIR2 <- 0}
if(!is.null(nrow(dfBAM_ASCNVLENGTH)) & !is.null(nrow(dfBAM_DECREASE))){reads_supporting_TADDIR2 <- reads_supporting_TADDIR2+nrow(dfBAM_ASCNVLENGTH)}
reads_supporting_TANDEM_DIRECT <- reads_supporting_TADDIR1 + reads_supporting_TADDIR2
#-----------------------------------------------------------------------------
## ACCESS BAM TANDEM DUPLICATIONS INDIRECT - reads orientation -/- or +/+ AND lower INSERT SIZE
# reads orientation -/-
flagTD_INDIR1 <- Rsamtools::scanBamFlag(isPaired = TRUE, isMinusStrand=TRUE, isMateMinusStrand=TRUE)
parametersTD_INDIR1 <- Rsamtools::ScanBamParam(what = c("pos", "qwidth","isize"), which=region, flag=flagTD_INDIR1)
dfBAM <- Rsamtools::scanBam(bamFile, param=parametersTD_INDIR1)[[1]]
dfBAM <- as.data.frame(dfBAM)
dfBAM <- na.omit(dfBAM)
# %% Circular genome correction
if(!is.null(nrow(dfBAM))){
# % positive values
REF <- referenceLength/2
dfBAM[dfBAM$isize>REF,"isize"] <- dfBAM[dfBAM$isize>REF,"isize"] - 2*dfBAM[dfBAM$isize>REF,"qwidth"] - referenceLength
# % negative values
REF <- referenceLength/-2
dfBAM[dfBAM$isize<REF,"isize"] <- dfBAM[dfBAM$isize<REF,"isize"] + 2*dfBAM[dfBAM$isize<REF,"qwidth"] + referenceLength
#find outliers in TLEN fragment size 1.5xIQR rule or as same as CNV length
out_ind <- which(abs(dfBAM$isize)>(CNVLENGTH-2*insertSize) & abs(dfBAM$isize)<(CNVLENGTH+2*insertSize))
dfBAM_ASCNVLENGTH<-dfBAM[out_ind,]
out_ind <- which(abs(dfBAM$isize)<(insertSize_1Q-1.5*insertSize_IQR))
dfBAM_DECREASE<-dfBAM[out_ind,]
}
if(!is.null(nrow(dfBAM_DECREASE))){reads_supporting_TADINDIR1 <- nrow(dfBAM_DECREASE)}else{reads_supporting_TADINDIR1 <- 0}
if(!is.null(nrow(dfBAM_ASCNVLENGTH)) & !is.null(nrow(dfBAM_DECREASE))){reads_supporting_TADINDIR1 <- reads_supporting_TADINDIR1+nrow(dfBAM_ASCNVLENGTH)}
# reads orientation +/+
flagTD_INDIR2 <- Rsamtools::scanBamFlag(isPaired = TRUE, isMinusStrand=FALSE, isMateMinusStrand=FALSE)
parametersTD_INDIR2 <- Rsamtools::ScanBamParam(what = c("pos", "qwidth","isize"), which=region, flag=flagTD_INDIR2)
dfBAM <- Rsamtools::scanBam(bamFile, param=parametersTD_INDIR2)[[1]]
dfBAM <- as.data.frame(dfBAM)
dfBAM <- na.omit(dfBAM)
# %% Circular genome correction
if(!is.null(nrow(dfBAM))){
# % positive values
REF <- referenceLength/2
dfBAM[dfBAM$isize>REF,"isize"] <- dfBAM[dfBAM$isize>REF,"isize"] - 2*dfBAM[dfBAM$isize>REF,"qwidth"] - referenceLength
# % negative values
REF <- referenceLength/-2
dfBAM[dfBAM$isize<REF,"isize"] <- dfBAM[dfBAM$isize<REF,"isize"] + 2*dfBAM[dfBAM$isize<REF,"qwidth"] + referenceLength
#find outliers in TLEN fragment size 1.5xIQR rule or as same as CNV length
out_ind <- which(abs(dfBAM$isize)>(CNVLENGTH-2*insertSize) & abs(dfBAM$isize)<(CNVLENGTH+2*insertSize))
dfBAM_ASCNVLENGTH<-dfBAM[out_ind,]
out_ind <- which(abs(dfBAM$isize)<(insertSize_1Q-1.5*insertSize_IQR))
dfBAM_DECREASE<-dfBAM[out_ind,]
}
if(!is.null(nrow(dfBAM_DECREASE))){reads_supporting_TADINDIR2 <- nrow(dfBAM_DECREASE)}else{reads_supporting_TADINDIR2 <- 0}
if(!is.null(nrow(dfBAM_ASCNVLENGTH)) & !is.null(nrow(dfBAM_DECREASE))){reads_supporting_TADINDIR2 <- reads_supporting_TADINDIR2+nrow(dfBAM_ASCNVLENGTH)}
reads_supporting_TANDEM_INDIRECT <- reads_supporting_TADINDIR1 + reads_supporting_TADINDIR2
################################################################################################################################################################
#-----------------------------------------------------------------------------
# ACCESS BAM INTERSPERSED DUPLICATIONS INVERSED - reads orientation -/- OR +/+ AND much higher INSERT SIZE
# reads orientation -/-
flagINTERDUP_INV1 <- Rsamtools::scanBamFlag(isPaired = TRUE, isMinusStrand=TRUE, isMateMinusStrand=TRUE)
parametersINTERDUP_INV1 <- Rsamtools::ScanBamParam(what = c("pos", "qwidth","isize"), which=region, flag=flagINTERDUP_INV1)
dfBAM <- Rsamtools::scanBam(bamFile, param=parametersINTERDUP_INV1)[[1]]
dfBAM <- as.data.frame(dfBAM)
dfBAM <- na.omit(dfBAM)
# %% Circular genome correction
if(!is.null(nrow(dfBAM))){
# % positive values
REF <- referenceLength/2
dfBAM[dfBAM$isize>REF,"isize"] <- dfBAM[dfBAM$isize>REF,"isize"] - 2*dfBAM[dfBAM$isize>REF,"qwidth"] - referenceLength
# % negative values
REF <- referenceLength/-2
dfBAM[dfBAM$isize<REF,"isize"] <- dfBAM[dfBAM$isize<REF,"isize"] + 2*dfBAM[dfBAM$isize<REF,"qwidth"] + referenceLength
#find outliers in TLEN fragment size 1.5xIQR rule
out_ind <- which(abs(dfBAM$isize)>(insertSize_3Q+10*insertSize_IQR))
dfBAM_INCREASE<-dfBAM[out_ind,]
}
if(!is.null(nrow(dfBAM_INCREASE))){reads_count_INTERDUP_INV1 <- nrow(dfBAM_INCREASE)}else{reads_count_INTERDUP_INV1 <- 0}
# reads orientation +/+
flagINTERDUP_INV2 <- Rsamtools::scanBamFlag(isPaired = TRUE, isMinusStrand=FALSE, isMateMinusStrand=FALSE)
parametersINTERDUP_INV2 <- Rsamtools::ScanBamParam(what = c("pos", "qwidth","isize"),which=region,flag=flagINTERDUP_INV2)
dfBAM <- Rsamtools::scanBam(bamFile, param=parametersINTERDUP_INV2)[[1]]
dfBAM <- as.data.frame(dfBAM)
dfBAM <- na.omit(dfBAM)
# %% Circular genome correction
if(!is.null(nrow(dfBAM))){
# % positive values
REF <- referenceLength/2
dfBAM[dfBAM$isize>REF,"isize"] <- dfBAM[dfBAM$isize>REF,"isize"] - 2*dfBAM[dfBAM$isize>REF,"qwidth"] - referenceLength
# % negative values
REF <- referenceLength/-2
dfBAM[dfBAM$isize<REF,"isize"] <- dfBAM[dfBAM$isize<REF,"isize"] + 2*dfBAM[dfBAM$isize<REF,"qwidth"] + referenceLength
#find outliers in TLEN fragment size 1.5xIQR rule
out_ind <- which(abs(dfBAM$isize)>(insertSize_3Q+10*insertSize_IQR))
dfBAM_INCREASE<-dfBAM[out_ind,]
}
if(!is.null(nrow(dfBAM_INCREASE))){reads_count_INTERDUP_INV2 <- nrow(dfBAM_INCREASE)}else{reads_count_INTERDUP_INV2 <- 0}
reads_supporting_INTERSPERSED_INDIRECT <- reads_count_INTERDUP_INV2 + reads_count_INTERDUP_INV1
#-----------------------------------------------------------------------------
# ACCESS BAM INTERSPERSED DUPLICATIONS DIRECT - reads orientation +/- OR -/+ AND much higher INSERT SIZE
# reads orientation +/-
flagINTERDUP_DIR1 <- Rsamtools::scanBamFlag(isPaired = TRUE, isMinusStrand=FALSE, isFirstMateRead=TRUE,isMateMinusStrand=TRUE)
parametersINTERDUP_DIR1 <- Rsamtools::ScanBamParam(what = c("pos", "qwidth","isize"), which=region, flag=flagINTERDUP_DIR1)
dfBAM <- Rsamtools::scanBam(bamFile, param=parametersINTERDUP_DIR1)[[1]]
dfBAM <- as.data.frame(dfBAM)
dfBAM <- na.omit(dfBAM)
# %% Circular genome correction
if(!is.null(nrow(dfBAM))){
# % positive values
REF <- referenceLength/2
dfBAM[dfBAM$isize>REF,"isize"] <- dfBAM[dfBAM$isize>REF,"isize"] - 2*dfBAM[dfBAM$isize>REF,"qwidth"] - referenceLength
# % negative values
REF <- referenceLength/-2
dfBAM[dfBAM$isize<REF,"isize"] <- dfBAM[dfBAM$isize<REF,"isize"] + 2*dfBAM[dfBAM$isize<REF,"qwidth"] + referenceLength
#find outliers in TLEN fragment size 1.5xIQR rule
out_ind <- which(abs(dfBAM$isize)>(insertSize_3Q+10*insertSize_IQR))
dfBAM_INCREASE<-dfBAM[out_ind,]
}
if(!is.null(nrow(dfBAM_INCREASE))){reads_count_INTERDUP_DIR1 <- nrow(dfBAM_INCREASE)}else{reads_count_INTERDUP_DIR1 <- 0}
# reads orientation -/+
flagINTERDUP_DIR2 <- Rsamtools::scanBamFlag(isPaired = TRUE, isMinusStrand=TRUE, isFirstMateRead=TRUE,isMateMinusStrand=FALSE)
parametersINTERDUP_DIR2 <- Rsamtools::ScanBamParam(what = c("pos", "qwidth","isize"),which=region,flag=flagINTERDUP_DIR2)
dfBAM <- Rsamtools::scanBam(bamFile, param=parametersINTERDUP_DIR2)[[1]]
dfBAM <- as.data.frame(dfBAM)
dfBAM <- na.omit(dfBAM)
# %% Circular genome correction
if(!is.null(nrow(dfBAM))){
# % positive values
REF <- referenceLength/2
dfBAM[dfBAM$isize>REF,"isize"] <- dfBAM[dfBAM$isize>REF,"isize"] - 2*dfBAM[dfBAM$isize>REF,"qwidth"] - referenceLength
# % negative values
REF <- referenceLength/-2
dfBAM[dfBAM$isize<REF,"isize"] <- dfBAM[dfBAM$isize<REF,"isize"] + 2*dfBAM[dfBAM$isize<REF,"qwidth"] + referenceLength
#find outliers in TLEN fragment size 1.5xIQR rule
out_ind <- which(abs(dfBAM$isize)>(insertSize_3Q+10*insertSize_IQR))
dfBAM_INCREASE<-dfBAM[out_ind,]
}
if(!is.null(nrow(dfBAM_INCREASE))){reads_count_INTERDUP_DIR2 <- nrow(dfBAM_INCREASE)}else{reads_count_INTERDUP_DIR2 <- 0}
reads_supporting_INTERSPERSED_DIRECT <- reads_count_INTERDUP_DIR2 + reads_count_INTERDUP_DIR1
#-----------------------------------------------------------------------------
# #print absolute numbers
CNV_DF[i,"reads_TOTAL"] <- reads_TOTAL
CNV_DF[i,"reads_supporting_Deletion"] <- reads_supporting_DELETION
CNV_DF[i,"reads_supporting_Tandem_Direct"] <- reads_supporting_TANDEM_DIRECT
CNV_DF[i,"reads_supporting_Tandem_Indirect"] <- reads_supporting_TANDEM_INDIRECT
CNV_DF[i,"reads_supporting_Interspersed_Direct"] <- reads_supporting_INTERSPERSED_DIRECT
CNV_DF[i,"reads_supporting_Interspersed_Indirect"] <- reads_supporting_INTERSPERSED_INDIRECT
#print Percentages
# CNV_DF[i,"reads_TOTAL"] <- reads_TOTAL
# CNV_DF[i,"reads_INSERT_INCREASE"] <- reads_count_INCREASE/(reads_TOTAL/100)
# CNV_DF[i,"reads_INSERT_DECREASE"] <- reads_count_DECREASE/(reads_TOTAL/100)
# CNV_DF[i,"reads_count_TAD_DUP"] <- reads_count_TD/(reads_TOTAL/100)
# CNV_DF[i,"reads_count_INTER_DUP_DIR"] <- reads_count_INTERDUP_DIR/(reads_TOTAL/100)
# CNV_DF[i,"reads_count_INTER_DUP_INV"] <- reads_count_INTERDUP_INV/(reads_TOTAL/100)
# SUBTYPE SELECTION
signaturesSUM <- reads_supporting_DELETION+reads_supporting_TANDEM_DIRECT+reads_supporting_TANDEM_INDIRECT+reads_supporting_INTERSPERSED_DIRECT+reads_supporting_INTERSPERSED_INDIRECT
tempX <- which.max(c(reads_supporting_TANDEM_DIRECT,reads_supporting_TANDEM_INDIRECT,reads_supporting_INTERSPERSED_DIRECT,reads_supporting_INTERSPERSED_INDIRECT))
if(CNV_DF[i,"TYPE"] == "DEL") {CNV_DF[i,"SUBTYPE"] <- "DEL"}
if(CNV_DF[i,"TYPE"] == "DUP" & tempX==1) {CNV_DF[i,"SUBTYPE"] <- "TANDEM_DIRECT"}
if(CNV_DF[i,"TYPE"] == "DUP" & tempX==2) {CNV_DF[i,"SUBTYPE"] <- "TANDEM_INDIRECT"}
if(CNV_DF[i,"TYPE"] == "DUP" & tempX==3) {CNV_DF[i,"SUBTYPE"] <- "INTERSPERSED_DIRECT"}
if(CNV_DF[i,"TYPE"] == "DUP" & tempX==4) {CNV_DF[i,"SUBTYPE"] <- "INTERSPERSED_INDIRECT"}
if(signaturesSUM < (reads_TOTAL/10) ) {CNV_DF[i,"SUBTYPE"] <- "FALSE POSITIVE"}
#reorder
CNV_DF <- CNV_DF[,c("ID",
"START",
"END",
"LENGTH",
"COVERAGE",
"TYPE",
"SUBTYPE",
"reads_TOTAL",
"reads_supporting_Deletion",
"reads_supporting_Tandem_Direct",
"reads_supporting_Tandem_Indirect",
"reads_supporting_Interspersed_Direct",
"reads_supporting_Interspersed_Indirect"
)]
}
} else{CNV_DF <- data.frame(ID=as.character(), START=integer(), END=integer(), LENGTH=integer(), COVERAGE=integer(), TYPE=character(),SUBTYPE=as.character(),
reads_TOTAL=integer(),reads_supporting_Deletion=integer(),reads_supporting_Tandem_Direct=integer(),reads_supporting_Tandem_Indirect=integer(),
reads_supporting_Interspersed_Direct=integer(),reads_supporting_Interspersed_Indirect=integer())
}
return(CNV_DF)
}
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