package core import org.broadinstitute.sting.queue.extensions.gatk._ import org.broadinstitute.sting.queue.extensions.picard.PicardBamFunction import org.broadinstitute.sting.queue.QScript import org.broadinstitute.sting.queue.function.ListWriterFunction import net.sf.samtools.{SAMFileReader,SAMReadGroupRecord} import scala.io.Source._ import collection.JavaConversions._ import org.broadinstitute.sting.gatk.walkers.indels.IndelRealigner.ConsensusDeterminationModel class DataProcessingPipeline extends QScript { qscript => /**************************************************************************** * Required Parameters (if default values are not good for you) ****************************************************************************/ @Input(doc="input BAM file - or list of BAM files", fullName="input", shortName="i", required=true) var input: File = _ @Input(doc="path to GenomeAnalysisTK.jar", fullName="path_to_gatk_jar", shortName="gatk", required=true) var GATKjar: File = _ @Input(doc="path to AnalyzeCovariates.jar", fullName="path_to_ac_jar", shortName="ac", required=true) var ACJar: File = _ @Input(doc="path to R resources folder inside the Sting repository", fullName="path_to_r", shortName="r", required=true) var R: String = _ @Input(doc="path to Picard's MarkDuplicates.jar", fullName="path_to_dedup_jar", shortName="dedup", required=false) var dedupJar: File = new File("/seq/software/picard/current/bin/MarkDuplicates.jar") @Input(doc="path to Picard's MergeSamFiles.jar", fullName="path_to_merge_jar", shortName="merge", required=false) var mergeBamJar: File = new File("/seq/software/picard/current/bin/MergeSamFiles.jar") @Input(doc="path to Picard's ValidateSamFile.jar", fullName="path_to_validate_jar", shortName="validate", required=false) var validateSamJar: File = new File("/seq/software/picard/current/bin/ValidateSamFile.jar") @Input(doc="Reference fasta file", fullName="reference", shortName="R", required=false) var reference: File = new File("/seq/references/Homo_sapiens_assembly19/v1/Homo_sapiens_assembly19.fasta") /**************************************************************************** * Optional Parameters ****************************************************************************/ @Input(doc="path to Picard's RevertSam.jar (if re-aligning a previously processed BAM file)", fullName="path_to_revert_jar", shortName="revert", required=false) var revertSamJar: File = _ @Input(doc="path to Picard's SortSam.jar (if re-aligning a previously processed BAM file)", fullName="path_to_sort_jar", shortName="sort", required=false) var sortSamJar: File = _ @Input(doc="The path to the binary of bwa (usually BAM files have already been mapped - but if you want to remap this is the option)", fullName="path_to_bwa", shortName="bwa", required=false) var bwaPath: File = _ @Input(doc="dbsnp ROD to use (must be in VCF format)", fullName="dbsnp", shortName="D", required=false) var dbSNP: File = new File("/humgen/gsa-hpprojects/GATK/data/dbsnp_132_b37.leftAligned.vcf") @Input(doc="extra VCF files to use as reference indels for Indel Realignment", fullName="extra_indels", shortName="indels", required=false) var indels: File = new File("/humgen/gsa-hpprojects/GATK/data/Comparisons/Unvalidated/AFR+EUR+ASN+1KG.dindel_august_release_merged_pilot1.20110126.sites.vcf") @Input(doc="the project name determines the final output (BAM file) base name. Example NA12878 yields NA12878.processed.bam", fullName="project", shortName="p", required=false) var projectName: String = "project" @Input(doc="Output path for the processed BAM files.", fullName="output_directory", shortName="outputDir", required=false) var outputDir: String = "" @Input(doc="the -L interval string to be used by GATK - output bams at interval only", fullName="gatk_interval_string", shortName="L", required=false) var intervalString: String = "" @Input(doc="an intervals file to be used by GATK - output bams at intervals only", fullName="gatk_interval_file", shortName="intervals", required=false) var intervals: File = _ @Input(doc="Cleaning model: KNOWNS_ONLY, USE_READS or USE_SW", fullName="clean_model", shortName="cm", required=false) var cleaningModel: String = "USE_READS" @Input(doc="Decompose input BAM file and fully realign it using BWA and assume Single Ended reads", fullName="use_bwa_single_ended", shortName="bwase", required=false) var useBWAse: Boolean = false @Input(doc="Decompose input BAM file and fully realign it using BWA and assume Pair Ended reads", fullName="use_bwa_pair_ended", shortName="bwape", required=false) var useBWApe: Boolean = false /**************************************************************************** * Global Variables ****************************************************************************/ val queueLogDir: String = ".qlog/" // Gracefully hide Queue's output var nContigs: Int = 0 // Use the number of contigs for scatter gathering jobs var cleanModelEnum: ConsensusDeterminationModel = ConsensusDeterminationModel.USE_READS if (cleaningModel == "KNOWNS_ONLY") { cleanModelEnum = ConsensusDeterminationModel.KNOWNS_ONLY } else if (cleaningModel == "USE_SW") { cleanModelEnum = ConsensusDeterminationModel.USE_SW } /**************************************************************************** * Helper classes and methods ****************************************************************************/ // Utility function to check if there are multiple samples in a BAM file (currently we can't deal with that) def hasMultipleSamples(readGroups: java.util.List[SAMReadGroupRecord]): Boolean = { var sample: String = "" for (r <- readGroups) { if (sample.isEmpty) sample = r.getSample else if (sample != r.getSample) return true; } return false } // Utility function to merge all bam files of similar samples. Generates on BAM file per sample. // It uses the sample information on the header of the input BAM files. // // Because the realignment only happens after these scripts are executed, in case you are using // bwa realignment, this function will operate over the original bam files and output over the // (to be realigned) bam files. def createSampleFiles(bamFiles: List[File], realignedBamFiles: List[File] = null): Map[String, File] = { assert(bamFiles.length == realignedBamFiles.length, "List of orignal files must have the same number of files than the list of realigned bam files: " + bamFiles.length + " / " + realignedBamFiles.length) // Creating a table with SAMPLE information from each input BAM file val sampleTable = scala.collection.mutable.Map.empty[String, List[File]] val realignedIterator = realignedBamFiles.iterator for (bam <- bamFiles) { val rBam = realignedIterator.next // advance to next element in the realignedBam list so they're in sync. val samReader = new SAMFileReader(bam) val header = samReader.getFileHeader val readGroups = header.getReadGroups // only allow one sample per file. Bam files with multiple samples would require pre-processing of the file // with PrintReads to separate the samples. Tell user to do it himself! assert(!hasMultipleSamples(readGroups), "The pipeline requires that only one sample is present in a BAM file. Please separate the samples in " + bam) // Fill out the sample table with the readgroups in this file for (rg <- readGroups) { val sample = rg.getSample if (!sampleTable.contains(sample)) sampleTable(sample) = List(rBam) else if ( !sampleTable(sample).contains(rBam)) sampleTable(sample) :+= rBam } } // Creating one file for each sample in the dataset val sampleBamFiles = scala.collection.mutable.Map.empty[String, File] for ((sample, flist) <- sampleTable) { val sampleFileName = new File(qscript.outputDir + qscript.projectName + "." + sample + ".bam") sampleBamFiles(sample) = sampleFileName add(joinBams(flist, sampleFileName)) } return sampleBamFiles.toMap } // Checks how many contigs are in the dataset. Uses the BAM file header information. def getNumberOfContigs(bamFile: File): Int = { val samReader = new SAMFileReader(new File(bamFile)) return samReader.getFileHeader.getSequenceDictionary.getSequences.size() } // Rebuilds the Read Group string to give BWA def buildReadGroupString(samReader: SAMFileReader): List[String] = { val readGroups = samReader.getFileHeader.getReadGroups var l: List[String] = List() for (rg <- readGroups) { l :+= "@RG\t" + SAMReadGroupRecord.READ_GROUP_ID_TAG + ":" + rg.getReadGroupId + "\t" + SAMReadGroupRecord.PLATFORM_TAG + ":" + rg.getPlatformUnit + "\t" + SAMReadGroupRecord.LIBRARY_TAG + ":" + rg.getLibrary + "\t" + SAMReadGroupRecord.READ_GROUP_SAMPLE_TAG + ":" + rg.getSample + "\t" + SAMReadGroupRecord.SEQUENCING_CENTER_TAG + ":" + rg.getSequencingCenter } return l } // Takes a list of processed BAM files and realign them using the BWA option requested (bwase or bwape). // Returns a list of realigned BAM files. def performAlignment(bams: List[File]): List[File] = { var realignedBams: List[File] = List() for (bam <- bams) { val saiFile1 = swapExt(bam, ".bam", "1.sai") val saiFile2 = swapExt(bam, ".bam", "2.sai") val realignedSamFile = swapExt(bam, ".bam", ".realigned.sam") val realignedBamFile = swapExt(bam, ".bam", ".realigned.bam") val readGroupString = buildReadGroupString(new SAMFileReader(bam)) if (useBWAse) { add(bwa_aln_se(bam, saiFile1), bwa_sam_se(bam, saiFile1, realignedSamFile, readGroupString)) } else { add(bwa_aln_pe(bam, saiFile1, 1), bwa_aln_pe(bam, saiFile2, 2), bwa_sam_pe(bam, saiFile1, saiFile2, realignedSamFile, readGroupString)) } add(sortSam(realignedSamFile, realignedBamFile)) realignedBams :+= realignedBamFile } return realignedBams } // Reads a BAM LIST file and creates a scala list with all the files def createListFromFile(in: File):List[File] = { if (in.toString.endsWith("bam")) return List(in) var l: List[File] = List() for (bam <- fromFile(in).getLines) l :+= new File(bam) return l } /**************************************************************************** * Main script ****************************************************************************/ def script = { // keep a record of the number of contigs in the first bam file in the list val bams = createListFromFile(input) nContigs = getNumberOfContigs(bams(0)) val realignedBams = if (useBWApe || useBWAse) {performAlignment(bams)} else {bams} // Generate a BAM file per sample joining all per lane files if necessary val sampleBamFiles: Map[String, File] = createSampleFiles(bams, realignedBams) println("nContigs: " + nContigs) // Final output list of processed bam files var cohortList: List[File] = List() // Simple progress report println("\nFound the following samples: ") for ((sample, file) <- sampleBamFiles) println("\t" + sample + " -> " + file) // If this is a 'knowns only' indel realignment run, do it only once for all samples. val globalIntervals = new File(outputDir + projectName + ".intervals") if (cleaningModel == ConsensusDeterminationModel.KNOWNS_ONLY) add(target(null, globalIntervals)) // Put each sample through the pipeline for ((sample, bam) <- sampleBamFiles) { // BAM files generated by the pipeline val cleanedBam = swapExt(bam, ".bam", ".clean.bam") val dedupedBam = swapExt(bam, ".bam", ".clean.dedup.bam") val recalBam = swapExt(bam, ".bam", ".clean.dedup.recal.bam") // Accessory files val targetIntervals = if (cleaningModel == ConsensusDeterminationModel.KNOWNS_ONLY) {globalIntervals} else {swapExt(bam, ".bam", ".intervals")} val metricsFile = swapExt(bam, ".bam", ".metrics") val preRecalFile = swapExt(bam, ".bam", ".pre_recal.csv") val postRecalFile = swapExt(bam, ".bam", ".post_recal.csv") val preOutPath = swapExt(bam, ".bam", ".pre") val postOutPath = swapExt(bam, ".bam", ".post") val preValidateLog = swapExt(bam, ".bam", ".pre.validation") val postValidateLog = swapExt(bam, ".bam", ".post.validation") add(validate(bam, preValidateLog)) if (cleaningModel != ConsensusDeterminationModel.KNOWNS_ONLY) add(target(bam, targetIntervals)) add(clean(bam, targetIntervals, cleanedBam), dedup(cleanedBam, dedupedBam, metricsFile), cov(dedupedBam, preRecalFile), recal(dedupedBam, preRecalFile, recalBam), cov(recalBam, postRecalFile), analyzeCovariates(preRecalFile, preOutPath), analyzeCovariates(postRecalFile, postOutPath), validate(recalBam, postValidateLog)) cohortList :+= recalBam } // output a BAM list with all the processed per sample files val cohortFile = new File(qscript.outputDir + qscript.projectName + ".cohort.list") add(writeList(cohortList, cohortFile)) } /**************************************************************************** * Classes (Walkers and non-GATK programs) ****************************************************************************/ // General arguments to GATK walkers trait CommandLineGATKArgs extends CommandLineGATK { this.jarFile = qscript.GATKjar this.reference_sequence = qscript.reference this.memoryLimit = 4 this.isIntermediate = true } case class target (inBams: File, outIntervals: File) extends RealignerTargetCreator with CommandLineGATKArgs { if (cleaningModel != ConsensusDeterminationModel.KNOWNS_ONLY) this.input_file :+= inBams this.out = outIntervals this.mismatchFraction = 0.0 this.rodBind :+= RodBind("dbsnp", "VCF", dbSNP) this.rodBind :+= RodBind("indels", "VCF", indels) this.scatterCount = nContigs this.analysisName = queueLogDir + outIntervals + ".target" this.jobName = queueLogDir + outIntervals + ".target" } case class clean (inBams: File, tIntervals: File, outBam: File) extends IndelRealigner with CommandLineGATKArgs { this.input_file :+= inBams this.targetIntervals = tIntervals this.out = outBam this.rodBind :+= RodBind("dbsnp", "VCF", dbSNP) this.rodBind :+= RodBind("indels", "VCF", qscript.indels) this.consensusDeterminationModel = consensusDeterminationModel this.compress = 0 this.scatterCount = nContigs this.analysisName = queueLogDir + outBam + ".clean" this.jobName = queueLogDir + outBam + ".clean" } case class cov (inBam: File, outRecalFile: File) extends CountCovariates with CommandLineGATKArgs { this.rodBind :+= RodBind("dbsnp", "VCF", dbSNP) this.covariate ++= List("ReadGroupCovariate", "QualityScoreCovariate", "CycleCovariate", "DinucCovariate") this.input_file :+= inBam this.recal_file = outRecalFile if (!qscript.intervalString.isEmpty()) this.intervalsString ++= List(qscript.intervalString) else if (qscript.intervals != null) this.intervals :+= qscript.intervals this.scatterCount = nContigs this.analysisName = queueLogDir + outRecalFile + ".covariates" this.jobName = queueLogDir + outRecalFile + ".covariates" } case class recal (inBam: File, inRecalFile: File, outBam: File) extends TableRecalibration with CommandLineGATKArgs { this.input_file :+= inBam this.recal_file = inRecalFile this.baq = org.broadinstitute.sting.utils.baq.BAQ.CalculationMode.CALCULATE_AS_NECESSARY this.out = outBam if (!qscript.intervalString.isEmpty()) this.intervalsString ++= List(qscript.intervalString) else if (qscript.intervals != null) this.intervals :+= qscript.intervals this.scatterCount = nContigs this.isIntermediate = false this.analysisName = queueLogDir + outBam + ".recalibration" this.jobName = queueLogDir + outBam + ".recalibration" } // Outside tools (not GATK walkers) case class analyzeCovariates (inRecalFile: File, outPath: File) extends AnalyzeCovariates { this.jarFile = qscript.ACJar this.resources = qscript.R this.recal_file = inRecalFile this.output_dir = outPath.toString this.analysisName = queueLogDir + inRecalFile + ".analyze_covariates" this.jobName = queueLogDir + inRecalFile + ".analyze_covariates" } case class dedup (inBam: File, outBam: File, metricsFile: File) extends PicardBamFunction { @Input(doc="fixed bam") var clean = inBam @Output(doc="deduped bam") var deduped = outBam @Output(doc="deduped bam index") var dedupedIndex = new File(outBam + "bai") @Output(doc="metrics file") var metrics = metricsFile override def inputBams = List(clean) override def outputBam = deduped override def commandLine = super.commandLine + " M=" + metricsFile this.sortOrder = null this.createIndex = true this.memoryLimit = 6 this.isIntermediate = true this.jarFile = qscript.dedupJar this.analysisName = queueLogDir + outBam + ".dedup" this.jobName = queueLogDir + outBam + ".dedup" } case class joinBams (inBams: List[File], outBam: File) extends PicardBamFunction { @Input(doc="input bam list") var join = inBams @Output(doc="joined bam") var joined = outBam @Output(doc="joined bam index") var joinedIndex = new File(outBam + "bai") override def inputBams = join override def outputBam = joined override def commandLine = super.commandLine + " CREATE_INDEX=true" this.jarFile = qscript.mergeBamJar this.isIntermediate = true this.analysisName = queueLogDir + outBam + ".joinBams" this.jobName = queueLogDir + outBam + ".joinBams" } case class sortSam (inSam: File, outBam: File) extends PicardBamFunction { @Input(doc="input unsorted sam file") var sam = inSam @Output(doc="sorted bam") var bam = outBam @Output(doc="sorted bam index") var bamIndex = new File(outBam + "bai") override def inputBams = List(sam) override def outputBam = bam override def commandLine = super.commandLine + " CREATE_INDEX=true" this.jarFile = qscript.sortSamJar this.isIntermediate = true this.analysisName = queueLogDir + outBam + ".sortSam" this.jobName = queueLogDir + outBam + ".sortSam" } case class validate (inBam: File, outLog: File) extends PicardBamFunction { @Input(doc="input bam list") var toValidate = inBam @Output(doc="validation log") var validate = outLog override def inputBams = List(inBam) override def outputBam = outLog override def commandLine = super.commandLine + " VALIDATE_INDEX=true MAX_RECORDS_IN_RAM=100000 MODE=SUMMARY REFERENCE_SEQUENCE=" + qscript.reference sortOrder = null this.jarFile = qscript.validateSamJar this.isIntermediate = false this.analysisName = queueLogDir + outLog + ".validate" this.jobName = queueLogDir + outLog + ".validate" } case class revert (inBam: File, outBam: File) extends PicardBamFunction { @Input(doc="old annotated bam") var oldBam = inBam @Output(doc="reverted bam") var revertedBam = outBam @Output(doc="reverted bam index") var revertedBamIndex = new File(outBam + ".bai") override def inputBams = List(oldBam) override def outputBam = revertedBam override def commandLine = super.commandLine + " CREATE_INDEX=true" this.isIntermediate = true this.jarFile = qscript.dedupJar this.analysisName = queueLogDir + outBam + ".dedup" this.jobName = queueLogDir + outBam + ".dedup" } case class bwa_aln_se (inBam: File, outSai: File) extends CommandLineFunction { @Input(doc="bam file to be aligned") var bam = inBam @Output(doc="output sai file") var sai = outSai def commandLine = bwaPath + " aln -q 5 " + reference + " -b " + bam + " > " + sai this.isIntermediate = true this.analysisName = queueLogDir + outSai + ".bwa_aln_se" this.jobName = queueLogDir + outSai + ".bwa_aln_se" } case class bwa_aln_pe (inBam: File, outSai1: File, index: Int) extends CommandLineFunction { @Input(doc="bam file to be aligned") var bam = inBam @Output(doc="output sai file for 1st mating pair") var sai = outSai1 def commandLine = bwaPath + " aln -q 5 " + reference + " -b" + index + " " + bam + " > " + sai this.isIntermediate = true this.analysisName = queueLogDir + outSai1 + ".bwa_aln_pe1" this.jobName = queueLogDir + outSai1 + ".bwa_aln_pe1" } case class bwa_sam_se (inBam: File, inSai: File, outBam: File, readGroup: List[String]) extends CommandLineFunction { @Input(doc="bam file to be aligned") var bam = inBam @Input(doc="bwa alignment index file") var sai = inSai @Output(doc="output aligned bam file") var alignedBam = outBam var readGroupParameters = "" for (rg <- readGroup) { readGroupParameters += " -r \'" + rg + "\'" } def commandLine = bwaPath + " samse " + readGroupParameters + " " + reference + " " + sai + " " + bam + " > " + alignedBam this.isIntermediate = true this.analysisName = queueLogDir + outBam + ".bwa_sam_se" this.jobName = queueLogDir + outBam + ".bwa_sam_se" } case class bwa_sam_pe (inBam: File, inSai1: File, inSai2:File, outBam: File, readGroup: List[String]) extends CommandLineFunction { @Input(doc="bam file to be aligned") var bam = inBam @Input(doc="bwa alignment index file for 1st mating pair") var sai1 = inSai1 @Input(doc="bwa alignment index file for 2nd mating pair") var sai2 = inSai2 @Output(doc="output aligned bam file") var alignedBam = outBam var readGroupParameters = "" for (rg <- readGroup) { readGroupParameters += " -r \'" + rg + "\'" } def commandLine = bwaPath + " sampe " + readGroupParameters + " " + reference + " " + sai1 + " " + sai2 + " " + bam + " " + bam + " > " + alignedBam this.isIntermediate = true this.analysisName = queueLogDir + outBam + ".bwa_sam_pe" this.jobName = queueLogDir + outBam + ".bwa_sam_pe" } case class writeList(inBams: List[File], outBamList: File) extends ListWriterFunction { this.inputFiles = inBams this.listFile = outBamList this.analysisName = queueLogDir + outBamList + ".bamList" this.jobName = queueLogDir + outBamList + ".bamList" } }