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package var2;
import java.io.PrintStream;
import java.util.ArrayList;
import fileIO.ByteFile;
import fileIO.FileFormat;
import fileIO.ReadWrite;
import shared.Parse;
import shared.Parser;
import shared.PreParser;
import shared.Shared;
import shared.Timer;
import shared.Tools;
import shared.TrimRead;
import stream.ConcurrentReadInputStream;
import stream.ConcurrentReadOutputStream;
import stream.FastaReadInputStream;
import stream.Read;
import stream.ReadStreamWriter;
import stream.SamLine;
import structures.ListNum;
import tracker.ReadStats;
/**
* Realigns samlines to a reference.
*
* @author Brian Bushnell
* @date April 26, 2017
*
*/
public class Realign {
/*--------------------------------------------------------------*/
/*---------------- Initialization ----------------*/
/*--------------------------------------------------------------*/
/**
* Code entrance from the command line.
* @param args Command line arguments
*/
public static void main(String[] args){
//Start a timer immediately upon code entrance.
Timer t=new Timer();
//Create an instance of this class
Realign x=new Realign(args);
//Run the object
x.process(t);
//Close the print stream if it was redirected
Shared.closeStream(x.outstream);
}
/**
* Constructor.
* @param args Command line arguments
*/
public Realign(String[] args){
{//Preparse block for help, config files, and outstream
PreParser pp=new PreParser(args, getClass(), false);
args=pp.args;
outstream=pp.outstream;
}
//Set static variables
SamLine.PARSE_OPTIONAL_MD_ONLY=true; //I only need the MD tag..
SamLine.RNAME_AS_BYTES=false;
SamLine.SET_FROM_OK=true;
ReadStreamWriter.USE_ATTACHED_SAMLINE=true;
ReadWrite.USE_PIGZ=ReadWrite.USE_UNPIGZ=true;
ReadWrite.setZipThreads(Shared.threads());
Shared.TRIM_READ_COMMENTS=Shared.TRIM_RNAME=true;
//Create a parser object
Parser parser=new Parser();
parser.qtrimLeft=qtrimLeft;
parser.qtrimRight=qtrimRight;
parser.trimq=trimq;
samFilter.includeUnmapped=false;
samFilter.includeSupplimentary=false;
samFilter.includeDuplicate=true;
samFilter.includeNonPrimary=true;
samFilter.includeQfail=false;
samFilter.minMapq=4;
//Parse each argument
for(int i=0; i<args.length; i++){
String arg=args[i];
//Break arguments into their constituent parts, in the form of "a=b"
String[] split=arg.split("=");
String a=split[0].toLowerCase();
String b=split.length>1 ? split[1] : null;
if(a.equals("verbose")){
verbose=Parse.parseBoolean(b);
}else if(a.equals("ordered")){
ordered=Parse.parseBoolean(b);
}else if(a.equals("parse_flag_goes_here")){
long fake_variable=Parse.parseKMG(b);
//Set a variable here
}
else if(a.equals("unclip")){
unclip=Parse.parseBoolean(b);
}else if(a.equals("realignrows") || a.equals("rerows")){
Realigner.defaultMaxrows=Integer.parseInt(b);
}else if(a.equals("realigncols") || a.equals("recols")){
Realigner.defaultColumns=Integer.parseInt(b);
}else if(a.equals("realignpadding") || a.equals("repadding") || a.equals("padding")){
Realigner.defaultPadding=Integer.parseInt(b);
}else if(a.equals("msa")){
Realigner.defaultMsaType=b;
}else if(a.equals("ref")){
ref=b;
}else if(a.equals("border")){
border=Integer.parseInt(b);
}
else if(samFilter.parse(arg, a, b)){
//do nothing
}
else if(parser.parse(arg, a, b)){//Parse standard flags in the parser
//do nothing
}else{
outstream.println("Unknown parameter "+args[i]);
assert(false) : "Unknown parameter "+args[i];
}
}
samFilter.setSamtoolsFilter();
{//Process parser fields
Parser.processQuality();
maxReads=parser.maxReads;
overwrite=ReadStats.overwrite=parser.overwrite;
append=ReadStats.append=parser.append;
in1=parser.in1;
out1=parser.out1;
trimq=parser.trimq;
trimE=parser.trimE();
extin=parser.extin;
extout=parser.extout;
}
assert(FastaReadInputStream.settingsOK());
//Ensure there is an input file
if(in1==null || ref==null){throw new RuntimeException("Error - one input file and one reference are required.");}
//Adjust the number of threads for input file reading
if(!ByteFile.FORCE_MODE_BF1 && !ByteFile.FORCE_MODE_BF2 && Shared.threads()>2){
ByteFile.FORCE_MODE_BF2=true;
}
//Ensure output files can be written
if(!Tools.testOutputFiles(overwrite, append, false, out1)){
outstream.println((out1==null)+", "+out1);
throw new RuntimeException("\n\noverwrite="+overwrite+"; Can't write to output files "+out1+"\n");
}
//Ensure input files can be read
if(!Tools.testInputFiles(false, true, in1, ref)){
throw new RuntimeException("\nCan't read some input files.\n");
}
//Ensure that no file was specified multiple times
if(!Tools.testForDuplicateFiles(true, in1, ref, out1)){
throw new RuntimeException("\nSome file names were specified multiple times.\n");
}
//Create output FileFormat objects
ffout1=FileFormat.testOutput(out1, FileFormat.SAM, extout, true, overwrite, append, ordered);
//Create input FileFormat objects
ffin1=FileFormat.testInput(in1, FileFormat.SAM, extin, true, true);
loadReference();
}
private void loadReference(){
if(loadedRef){return;}
assert(ref!=null);
ScafMap.loadReference(ref, scafMap, samFilter, true);
Realigner.map=scafMap;
loadedRef=true;
}
/*--------------------------------------------------------------*/
/*---------------- Outer Methods ----------------*/
/*--------------------------------------------------------------*/
/** Create read streams and process all data */
void process(Timer t){
//Turn off read validation in the input threads to increase speed
final boolean vic=Read.VALIDATE_IN_CONSTRUCTOR;
Read.VALIDATE_IN_CONSTRUCTOR=Shared.threads()<4;
//Create a read input stream
final ConcurrentReadInputStream cris;
{
cris=ConcurrentReadInputStream.getReadInputStream(maxReads, true, ffin1, null, null, null);
cris.start(); //Start the stream
if(verbose){outstream.println("Started cris");}
}
boolean paired=cris.paired();
if(!ffin1.samOrBam()){outstream.println("Input is being processed as "+(paired ? "paired" : "unpaired"));}
//Optionally create a read output stream
final ConcurrentReadOutputStream ros;
if(ffout1!=null){
//Select output buffer size based on whether it needs to be ordered
final int buff=(ordered ? Tools.mid(16, 128, (Shared.threads()*2)/3) : 8);
//Notify user of output mode
if(cris.paired() && (in1!=null && !ffin1.samOrBam() && !ffout1.samOrBam())){
outstream.println("Writing interleaved.");
}
ros=ConcurrentReadOutputStream.getStream(ffout1, null, null, null, buff, null, true);
ros.start(); //Start the stream
}else{ros=null;}
//Reset counters
readsProcessed=0;
basesProcessed=0;
//Process the reads in separate threads
spawnThreads(cris, ros);
if(verbose){outstream.println("Finished; closing streams.");}
//Write anything that was accumulated by ReadStats
errorState|=ReadStats.writeAll();
//Close the read streams
errorState|=ReadWrite.closeStreams(cris, ros);
//Reset read validation
Read.VALIDATE_IN_CONSTRUCTOR=vic;
//Report timing and results
t.stop();
outstream.println("Realignments: \t"+realignmentsAttempted);
outstream.println("Successes: \t"+realignmentsSucceeded);
outstream.println("Improvements: \t"+realignmentsImproved);
outstream.println("Retained: \t"+realignmentsRetained);
outstream.println("Bases trimmed: \t"+basesTrimmed);
outstream.println();
outstream.println(Tools.timeReadsBasesProcessed(t, readsProcessed, basesProcessed, 8));
//Throw an exception of there was an error in a thread
if(errorState){
throw new RuntimeException(getClass().getName()+" terminated in an error state; the output may be corrupt.");
}
}
/** Spawn process threads */
private void spawnThreads(final ConcurrentReadInputStream cris, final ConcurrentReadOutputStream ros){
//Do anything necessary prior to processing
//Determine how many threads may be used
final int threads=Shared.threads();
//Fill a list with ProcessThreads
ArrayList<ProcessThread> alpt=new ArrayList<ProcessThread>(threads);
for(int i=0; i<threads; i++){
alpt.add(new ProcessThread(cris, ros, i));
}
//Start the threads
for(ProcessThread pt : alpt){
pt.start();
}
//Wait for completion of all threads
boolean success=true;
for(ProcessThread pt : alpt){
//Wait until this thread has terminated
while(pt.getState()!=Thread.State.TERMINATED){
try {
//Attempt a join operation
pt.join();
} catch (InterruptedException e) {
//Potentially handle this, if it is expected to occur
e.printStackTrace();
}
}
//Accumulate per-thread statistics
readsProcessed+=pt.readsProcessedT;
basesProcessed+=pt.basesProcessedT;
trimmedBasesProcessed+=pt.trimmedBasesProcessedT;
basesTrimmed+=pt.basesTrimmedT;
readsDiscarded+=pt.readsDiscardedT;
pairedInSequencingReadsProcessed+=pt.pairedInSequencingReadsProcessedT;
properlyPairedReadsProcessed+=pt.properlyPairedReadsProcessedT;
realignmentsAttempted+=pt.realigner.realignmentsAttempted;
realignmentsImproved+=pt.realigner.realignmentsImproved;
realignmentsSucceeded+=pt.realigner.realignmentsSucceeded;
realignmentsRetained+=pt.realigner.realignmentsRetained;
success&=pt.success;
}
//Track whether any threads failed
if(!success){errorState=true;}
//Do anything necessary after processing
}
/*--------------------------------------------------------------*/
/*---------------- Inner Methods ----------------*/
/*--------------------------------------------------------------*/
/*--------------------------------------------------------------*/
/*---------------- Inner Classes ----------------*/
/*--------------------------------------------------------------*/
/** This class is static to prevent accidental writing to shared variables.
* It is safe to remove the static modifier. */
private class ProcessThread extends Thread {
//Constructor
ProcessThread(final ConcurrentReadInputStream cris_, final ConcurrentReadOutputStream ros_, final int tid_){
cris=cris_;
ros=ros_;
tid=tid_;
realigner=new Realigner();
}
//Called by start()
@Override
public void run(){
//Do anything necessary prior to processing
//Process the reads
processInner();
//Do anything necessary after processing
//Indicate successful exit status
success=true;
}
/** Iterate through the reads */
void processInner(){
//Grab the first ListNum of reads
ListNum<Read> ln=cris.nextList();
//Grab the actual read list from the ListNum
ArrayList<Read> reads=(ln!=null ? ln.list : null);
//As long as there is a nonempty read list...
while(ln!=null && reads!=null && reads.size()>0){//ln!=null prevents a compiler potential null access warning
// if(verbose){outstream.println("Fetched "+reads.size()+" reads.");} //Disabled due to non-static access
//Loop through each read in the list
for(int idx=0; idx<reads.size(); idx++){
final Read r1=reads.get(idx);
//Validate reads in worker threads
if(!r1.validated()){r1.validate(true);}
//Track the initial length for statistics
final int initialLength1=r1.length();
final int initialLength2=r1.mateLength();
//Increment counters
readsProcessedT+=r1.pairCount();
basesProcessedT+=initialLength1+initialLength2;
{
//Reads are processed in this block.
boolean keep=processRead(r1);
if(!keep){
reads.set(idx, null);
readsDiscardedT++;
}
}
}
//Output reads to the output stream
if(ros!=null){ros.add(reads, ln.id);}
//Notify the input stream that the list was used
cris.returnList(ln);
// if(verbose){outstream.println("Returned a list.");} //Disabled due to non-static access
//Fetch a new list
ln=cris.nextList();
reads=(ln!=null ? ln.list : null);
}
//Notify the input stream that the final list was used
if(ln!=null){
cris.returnList(ln.id, ln.list==null || ln.list.isEmpty());
}
}
/**
* Process a read.
* @param r Read 1
* @return True if the reads should be kept, false if they should be discarded.
*/
boolean processRead(final Read r){
if(r.bases==null || r.length()<=1){return false;}
final SamLine sl=r.samline;
final int len0=r.length();
if(samFilter!=null && !samFilter.passesFilter(sl)){return false;}
if(sl.properPair()){properlyPairedReadsProcessedT++;}
if(sl.hasMate()){pairedInSequencingReadsProcessedT++;}
final Scaffold scaf=scafMap.getScaffold(sl);
final int scafnum=scaf.number;
// r.toLongMatchString(false); //Not necessary if scoring can be done on short match string
assert(sl.cigar!=null) : sl;
boolean realigned=realigner.realign(r, sl, scaf, unclip);
if(!realigned && r.match!=null){sl.cigar=SamLine.toCigar14(r.match, r.start, r.stop, scaf.length, sl.seq);}
assert(sl.cigar!=null) : sl;
int leftTrimAmount=border, rightTrimAmount=border;
if(qtrimLeft || qtrimRight){
long packed=TrimRead.testOptimal(r.bases, r.quality, trimE);
if(qtrimLeft){leftTrimAmount=Tools.max(leftTrimAmount, (int)((packed>>32)&0xFFFFFFFFL));}
if(qtrimRight){rightTrimAmount=Tools.max(rightTrimAmount, (int)((packed)&0xFFFFFFFFL));}
}
if(len0-leftTrimAmount-rightTrimAmount<2){return false;}
int trimmed=(leftTrimAmount<1 && rightTrimAmount<1 ? 0 : TrimRead.trimReadWithMatch(r, sl, leftTrimAmount, rightTrimAmount, 0, scaf.length, false));
if(trimmed<0 || r.length()<2 || sl.cigar==null){return false;}
basesTrimmedT+=trimmed;
if(trimmed>0){sl.optional=null;}
assert(sl.cigar!=null) : leftTrimAmount+", "+rightTrimAmount+", "+len0+"\n"+sl;
return true;
}
/** Number of reads processed by this thread */
protected long readsProcessedT=0;
/** Number of bases processed by this thread */
protected long basesProcessedT=0;
/** Number of trimmed, mapped bases processed. */
protected long trimmedBasesProcessedT=0;
/** Number of trimmed, mapped bases processed. */
protected long basesTrimmedT=0;
/** Number of reads discarded by this thread */
protected long readsDiscardedT=0;
/** Number of paired reads processed by this thread, whether or not they mapped as pairs */
protected long pairedInSequencingReadsProcessedT=0;
/** Number of properly paired reads processed by this thread */
protected long properlyPairedReadsProcessedT=0;
/** True only if this thread has completed successfully */
boolean success=false;
/** Shared input stream */
private final ConcurrentReadInputStream cris;
/** Shared output stream */
private final ConcurrentReadOutputStream ros;
/** For realigning reads */
final Realigner realigner;
/** Thread ID */
final int tid;
}
/*--------------------------------------------------------------*/
/*---------------- Fields ----------------*/
/*--------------------------------------------------------------*/
/** Primary input file path */
private String in1=null;
/** A fasta file. */
private String ref=null;
/** Primary output file path */
private String out1=null;
/** Override input file extension */
private String extin=null;
/** Override output file extension */
private String extout=null;
/*--------------------------------------------------------------*/
/** Number of reads processed */
protected long readsProcessed=0;
/** Number of bases processed */
protected long basesProcessed=0;
/** Quit after processing this many input reads; -1 means no limit */
private long maxReads=-1;
/*--------------------------------------------------------------*/
private boolean loadedRef=false;
private boolean qtrimLeft=false;
private boolean qtrimRight=true;
private float trimq=10;
private final float trimE;
/*--------------------------------------------------------------*/
/** Number of trimmed, mapped bases processed */
protected long trimmedBasesProcessed=0;
/** Number of reads discarded */
protected long readsDiscarded=0;
/** Number of paired reads processed by this thread, whether or not they mapped as pairs */
protected long pairedInSequencingReadsProcessed=0;
/** Number of properly paired reads processed */
protected long properlyPairedReadsProcessed=0;
/** Number of bases trimmed */
protected long basesTrimmed=0;
protected long realignmentsAttempted;
protected long realignmentsImproved;
protected long realignmentsSucceeded;
protected long realignmentsRetained;
public final ScafMap scafMap=new ScafMap();
public int border=0;
public boolean unclip=false;
public final SamFilter samFilter=new SamFilter();
/*--------------------------------------------------------------*/
/*---------------- Final Fields ----------------*/
/*--------------------------------------------------------------*/
/** Primary input file */
private final FileFormat ffin1;
/** Primary output file */
private final FileFormat ffout1;
/*--------------------------------------------------------------*/
/*---------------- Common Fields ----------------*/
/*--------------------------------------------------------------*/
/** Print status messages to this output stream */
private PrintStream outstream=System.err;
/** Print verbose messages */
public static boolean verbose=false;
/** True if an error was encountered */
public boolean errorState=false;
/** Overwrite existing output files */
private boolean overwrite=true;
/** Append to existing output files */
private boolean append=false;
/** Reads are output in input order */
private boolean ordered=true;
}
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