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package jgi;
import java.io.File;
import java.io.PrintStream;
import java.util.ArrayList;
import java.util.Arrays;
import java.util.BitSet;
import java.util.HashMap;
import align2.QualityTools;
import dna.ChromosomeArray;
import dna.Data;
import dna.Scaffold;
import fileIO.ByteFile;
import fileIO.ByteStreamWriter;
import fileIO.FileFormat;
import fileIO.ReadWrite;
import fileIO.TextFile;
import fileIO.TextStreamWriter;
import shared.KillSwitch;
import shared.LineParser1;
import shared.Parse;
import shared.Parser;
import shared.PreParser;
import shared.Shared;
import shared.Timer;
import shared.Tools;
import shared.TrimRead;
import stream.Read;
import stream.SamLine;
import stream.SamLineStreamer;
import stream.ScaffoldCoordinates;
import stream.SiteScore;
import structures.ByteBuilder;
import structures.CoverageArray;
import structures.CoverageArray2;
import structures.CoverageArray3;
import structures.ListNum;
import structures.LongList;
import tracker.ReadStats;
/**
* @author Brian Bushnell
* @date Jan 4, 2013
*
*/
public class CoveragePileup {
/*--------------------------------------------------------------*/
/*---------------- Main ----------------*/
/*--------------------------------------------------------------*/
public static void main(String[] args){
Timer t=new Timer();
CoveragePileup x=new CoveragePileup(args);
x.process();
t.stop();
if(!KEY_VALUE){
x.outstream.println();
x.outstream.println("Time: \t"+t);
}
//Close the print stream if it was redirected
Shared.closeStream(x.outstream);
}
/*--------------------------------------------------------------*/
/*---------------- Initialization ----------------*/
/*--------------------------------------------------------------*/
public CoveragePileup(String[] args){
{//Preparse block for help, config files, and outstream
PreParser pp=new PreParser(args, printCommand ? getClass() : null, false);
args=pp.args;
outstream=pp.outstream;
}
int vectorMode=-1;
boolean outset=false;
ReadWrite.USE_UNPIGZ=true;
// SamLine.RNAME_AS_BYTES=false;
for(int i=0; i<args.length; i++){
final String arg=args[i];
final String[] split=arg.split("=");
String a=split[0].toLowerCase();
String b=split.length>1 ? split[1] : null;
if(a.equals("ref") || a.equals("reference") || a.equals("fasta")){
reference=b;
}else if(a.equals("addfromref")){
ADD_FROM_REF=Parse.parseBoolean(b);
}else if(a.equals("addfromreads")){
ADD_FROM_READS=Parse.parseBoolean(b);
}else if(a.equals("in") || a.equals("in1")){
in1=b;
}else if(a.equals("in2")){
in2=b;
}else if(a.equals("covin") || a.equals("incov")){
covIn.clear();
if(b!=null) {
for(String cf : Tools.commaPattern.split(b)) {covIn.add(cf);}
}
}else if(a.equals("out") || a.equals("coveragestats") || a.equals("covstats") || a.equals("stats")){
covstats=b;
outset=true;
}else if(a.equals("minscaf") || a.equals("covminscaf")){
minscaf=Integer.parseInt(b);
}else if(a.equals("mindepth") || a.equals("mincov")){
minDepthToBeCovered=Integer.parseInt(b);
}else if(a.equals("border")){
border=Integer.parseInt(b);
}else if(a.equals("qtrim")/* || a.equals("trim")*/){
if(b==null || b.length()==0){qtrimRight=qtrimLeft=true;}
else if(b.equalsIgnoreCase("left") || b.equalsIgnoreCase("l")){qtrimLeft=true;qtrimRight=false;}
else if(b.equalsIgnoreCase("right") || b.equalsIgnoreCase("r")){qtrimLeft=false;qtrimRight=true;}
else if(b.equalsIgnoreCase("both") || b.equalsIgnoreCase("rl") || b.equalsIgnoreCase("lr")){qtrimLeft=qtrimRight=true;}
else if(Tools.isDigit(b.charAt(0))){
trimq=Float.parseFloat(b);
qtrimRight=trimq>0;
}else{qtrimRight=qtrimLeft=Parse.parseBoolean(b);}
}else if(a.equals("trimq") || a.equals("trimquality")){
trimq=Float.parseFloat(b);
}else if(a.equals("ss") || a.equals("samstreamer")){
if(b!=null && Tools.isDigit(b.charAt(0))){
useStreamer=true;
streamerThreads=Tools.max(1, Integer.parseInt(b));
}else{
useStreamer=Parse.parseBoolean(b);
}
}else if(a.equals("minq") || a.equals("minmapq")){
minMapq=Integer.parseInt(b);
}else if(a.equals("outsam")){
outsam=b;
}else if(a.equals("rpkm") || a.equals("fpkm") || a.equals("outrpkm")){
outrpkm=b;
}else if(a.equals("outorf")){
outorf=b;
}else if(a.equals("orffasta") || a.equals("fastaorf")){
orffasta=b;
}else if(a.equals("basecov") || a.equals("outcov")){
basecov=b;
}else if(a.equals("ranges") || a.equals("rangecov")){
rangecov=b;
// }else if(a.equals("mincov") || a.equals("mindepth")){
// mincov=Integer.parseInt(b);
}else if(a.equals("bincov") || a.equals("outbinned")){
bincov=b;
}else if(a.equals("normcov") || a.equals("outnormalized")){
normcov=b;
}else if(a.equals("normcovo") || a.equals("outnormalizedoverall")){
normcovOverall=b;
}else if(a.equals("delta")){
DELTA_ONLY=Parse.parseBoolean(b);
}else if(a.equals("physical") || a.equals("physicalcoverage") || a.equals("physcov")){
PHYSICAL_COVERAGE=Parse.parseBoolean(b);
}else if(a.equals("tlen")){
USE_TLEN=Parse.parseBoolean(b);
}else if(a.equals("hist") || a.equals("histogram") || a.equals("covhist")){
histogram=b;
}else if(a.equals("reads")){
maxReads=Parse.parseKMG(b);
}else if(a.equals("scafs") || a.equals("scaffolds")){
initialScaffolds=Tools.max(128, (int)(Tools.min(Long.parseLong(b),2000000000)));
}else if(a.equals("binsize")){
binsize=Integer.parseInt(b);
}else if(a.equals("32bit")){
bits32=Parse.parseBoolean(b);
}else if(a.equals("bitset") || a.equals("usebitset") || a.equals("bitsets") || a.equals("usebitsets")){
// if(Parse.parseBoolean(b)){arrayMode=BITSET_MODE;}
vectorMode=Parse.parseBoolean(b) ? BITSET_MODE : NOTHING_MODE;
}else if(a.equals("array") || a.equals("arrays") || a.equals("usearrays")){
vectorMode=Parse.parseBoolean(b) ? ARRAY_MODE : NOTHING_MODE;
}else if(a.equals("median") || a.equals("calcmedian")){
if(Parse.parseBoolean(b)){
vectorMode=ARRAY_MODE;
}
}else if(a.startsWith("nonzero") || a.equals("nzo")){
NONZERO_ONLY=Parse.parseBoolean(b);
if(verbose){outstream.println("Set NONZERO_ONLY to "+NONZERO_ONLY);}
}else if(a.equals("append") || a.equals("app")){
append=ReadStats.append=Parse.parseBoolean(b);
}else if(a.equals("overwrite") || a.equals("ow")){
overwrite=Parse.parseBoolean(b);
if(verbose){outstream.println("Set overwrite to "+overwrite);}
}else if(a.equalsIgnoreCase("twocolumn")){
TWOCOLUMN=Parse.parseBoolean(b);
if(verbose){outstream.println("Set TWOCOLUMN to "+TWOCOLUMN);}
}else if(a.equalsIgnoreCase("keyvalue") || a.equalsIgnoreCase("machineout")){
KEY_VALUE=Parse.parseBoolean(b);
}else if(a.equalsIgnoreCase("countgc")){
COUNT_GC=Parse.parseBoolean(b);
if(verbose){outstream.println("Set COUNT_GC to "+COUNT_GC);}
}else if(a.equals("secondary") || a.equals("usesecondary")){
USE_SECONDARY=Parse.parseBoolean(b);
if(verbose){outstream.println("Set USE_SECONDARY_ALIGNMENTS to "+USE_SECONDARY);}
}else if(a.equals("softclip") || a.equals("includesoftclip")){
INCLUDE_SOFT_CLIP=Parse.parseBoolean(b);
if(verbose){outstream.println("Set INCLUDE_SOFT_CLIP to "+INCLUDE_SOFT_CLIP);}
}else if(a.equals("keepshortbins") || a.equals("ksb")){
KEEP_SHORT_BINS=Parse.parseBoolean(b);
if(verbose){outstream.println("Set KEEP_SHORT_BINS to "+KEEP_SHORT_BINS);}
}else if(a.equals("strandedcoverage") || a.equals("strandedcov") || a.equals("covstranded") || a.equals("stranded")){
STRANDED=Parse.parseBoolean(b);
}else if(a.equals("startcov") || a.equals("covstart") || a.equals("startonly")){
START_ONLY=Parse.parseBoolean(b);
}else if(a.equals("stopcov") || a.equals("covstop") || a.equals("stoponly")){
STOP_ONLY=Parse.parseBoolean(b);
}else if(a.equals("concise")){
CONCISE=Parse.parseBoolean(b);
}else if(a.equals("verbose")){
verbose=Parse.parseBoolean(b);
}else if(a.equals("normc") || a.equals("normalizecoverage")){
NORMALIZE_COVERAGE=Parse.parseBoolean(b);
}else if(a.equals("header") || a.equals("hdr")){
printHeader=Parse.parseBoolean(b);
}else if(a.equals("headerpound") || a.equals("#")){
headerPound=Parse.parseBoolean(b);
}else if(a.equals("stdev")){
calcCovStdev=Parse.parseBoolean(b);
}else if(a.equals("delcov") || a.equals("dels") || a.equals("includedels") || a.equals("includedeletions") || a.equals("delcoverage")){
INCLUDE_DELETIONS=Parse.parseBoolean(b);
}else if(a.equals("dupecoverage") || a.equals("dupecov") || a.equals("dupes") || a.equals("duplicates") || a.equals("includeduplicates")){
INCLUDE_DUPLICATES=Parse.parseBoolean(b);
}else if(a.equals("ignoredupes") || a.equals("ignoreduplicates")){
INCLUDE_DUPLICATES=!Parse.parseBoolean(b);
}else if(a.equals("normb") || a.equals("normalizebins")){
try {
NORMALIZE_LENGTH_BINS=Integer.parseInt(b);
} catch (NumberFormatException e) {
boolean x=Parse.parseBoolean(b);
NORMALIZE_LENGTH_BINS=x ? 100 : -1;
}
}else if(a.equals("covwindow")){
if(b==null || b.length()<1 || Character.isLetter(b.charAt(0))){
USE_WINDOW=Parse.parseBoolean(b);
}else{
LOW_COV_WINDOW=Integer.parseInt(b);
USE_WINDOW=(LOW_COV_WINDOW>0);
}
}else if(a.equals("covwindowavg")){
LOW_COV_DEPTH=Double.parseDouble(b);
}else if(a.equals("k")){
k=Integer.parseInt(b);
}else if(Parser.parseCommonStatic(arg, a, b)){
//do nothing
}else if(Parser.parseZip(arg, a, b)){
//do nothing
}else if(in1==null && arg.indexOf('=')<0 && new File(arg).exists()){
in1=arg;
}else{
throw new RuntimeException("Unknown parameter: "+args[i]);
}
}
trimE=(float)QualityTools.phredToProbError(trimq);
// assert(false) : qtrimLeft+", "+qtrimRight+", "+trimq+", "+trimE;
ReadWrite.SAMTOOLS_IGNORE_FLAG|=ReadWrite.SAM_UNMAPPED;
if(!USE_SECONDARY){ReadWrite.SAMTOOLS_IGNORE_FLAG|=ReadWrite.SAM_SECONDARY;}
if(!INCLUDE_DUPLICATES){ReadWrite.SAMTOOLS_IGNORE_FLAG|=ReadWrite.SAM_DUPLICATE;}
if(outsam==null){
SamLine.PARSE_0=false;
SamLine.PARSE_6=false;
SamLine.PARSE_7=false;
SamLine.PARSE_8=false;
if(k<1 && trimq<1){SamLine.PARSE_10=false;}
SamLine.PARSE_OPTIONAL=false;
}
if(vectorMode>-1){
USE_BITSETS=(vectorMode==BITSET_MODE);
USE_COVERAGE_ARRAYS=(vectorMode==ARRAY_MODE);
}else{
if(histogram==null && basecov==null && bincov==null && normcov==null && rangecov==null &&
normcovOverall==null && outorf==null && !calcCovStdev){//No need for coverage array!
USE_COVERAGE_ARRAYS=false;
if(TWOCOLUMN){//No need for bitset, either!
USE_BITSETS=false;
}else{
USE_BITSETS=true;
}
}
}
if(verbose){
outstream.println("Set USE_COVERAGE_ARRAYS to "+USE_COVERAGE_ARRAYS);
outstream.println("Set USE_BITSETS to "+USE_BITSETS);
}
if(maxReads<0){maxReads=Long.MAX_VALUE;}
{
final String a=(args.length>0 ? args[0] : null);
// final String b=(args.length>1 ? args[1] : null);
if(in1==null && a!=null && a.indexOf('=')<0 && (a.startsWith("stdin") || new File(a).exists())){in1=a;}
// if(covstats==null && b!=null && b.indexOf('=')<0){covstats=b;}
if(in1==null){in1="stdin";}
// if(covstats==null && !outset){
//// out="stdout";
//// outstream.println("Warning: output destination not set; producing no output. To print to standard out, set 'out=stdout'");
// outstream=System.err;
// }
}
assert(in1!=null);
// assert(out!=null || outset) : "Output file was not set.";
if(STRANDED){
assert(basecov==null || basecov.indexOf('#')>=0) : "Output filenames must contain '#' symbol for strand-specific output.";
assert(bincov==null || bincov.indexOf('#')>=0) : "Output filenames must contain '#' symbol for strand-specific output.";
assert(normcov==null || normcov.indexOf('#')>=0) : "Output filenames must contain '#' symbol for strand-specific output.";
assert(normcovOverall==null || normcovOverall.indexOf('#')>=0) : "Output filenames must contain '#' symbol for strand-specific output.";
assert(histogram==null || histogram.indexOf('#')>=0) : "Output filenames must contain '#' symbol for strand-specific output.";
assert(covstats==null || covstats.indexOf('#')>=0) : "Output filenames must contain '#' symbol for strand-specific output.";
}
if(!Tools.testInputFiles(false, true, in1, in2, reference)){
throw new RuntimeException("\nCan't read some input files.\n");
}
if(!Tools.testOutputFiles(overwrite, append, false, basecov, rangecov, bincov, normcov, normcovOverall, histogram, covstats, outrpkm)){
throw new RuntimeException("\n\noverwrite="+overwrite+"; Can't write to output files "+
basecov+", "+rangecov+", "+bincov+", "+normcov+", "+normcovOverall+", "+histogram+", "+covstats+", "+outrpkm+"\n");
}
}
/*--------------------------------------------------------------*/
/*---------------- Data Structures ----------------*/
/*--------------------------------------------------------------*/
/** The goal if this is to garbage-collect unnecessary objects, not really for reusing the object */
public void clear(){
list=null;
table=null;
pairTable=null;
program=null;
version=null;
in1=null;
covstats=null;
outsam=null;
outorf=null;
outrpkm=null;
reference=null;
histogram=null;
basecov=null;
bincov=null;
normcov=null;
normcovOverall=null;
orffasta=null;
error=false;
refBases=0;
mappedBases=0;
mappedNonClippedBases=0;
mappedBasesWithDels=0;
mappedReads=0;
properPairs=0;
readsProcessed=0;
basesProcessed=0;
kmersProcessed=0;
mappedKmers=0;
totalCoveredBases1=0;
totalCoveredBases2=0;
scaffoldsWithCoverage1=0;
scaffoldsWithCoverage2=0;
totalScaffolds=0;
}
public void createDataStructures(){
refBases=0;
mappedBases=0;
mappedNonClippedBases=0;
mappedBasesWithDels=0;
mappedReads=0;
properPairs=0;
readsProcessed=0;
basesProcessed=0;
kmersProcessed=0;
mappedKmers=0;
totalCoveredBases1=0;
totalCoveredBases2=0;
scaffoldsWithCoverage1=0;
scaffoldsWithCoverage2=0;
totalScaffolds=0;
error=false;
list=new ArrayList<Scaffold>(initialScaffolds);
table=new HashMap<String, Scaffold>(initialScaffolds);
if(PHYSICAL_COVERAGE){
pairTable=new HashMap<String, SamLine>();
if(COUNT_GC){
COUNT_GC=false;
outstream.println("COUNT_GC disabled for physical coverage mode.");
}
if(USE_SECONDARY){
USE_SECONDARY=false;
outstream.println("USE_SECONDARY disabled for physical coverage mode.");
}
SamLine.PARSE_0=true;
SamLine.PARSE_6=true;
SamLine.PARSE_7=true;
SamLine.PARSE_8=true;
SamLine.PARSE_10=false;
SamLine.PARSE_OPTIONAL=false;
}
}
/*--------------------------------------------------------------*/
/*---------------- Outer Methods ----------------*/
/*--------------------------------------------------------------*/
/** Read and process all input data. */
public void process(){
createDataStructures();
// System.err.println("A");
if(in1!=null && FileFormat.isBamFile(in1)){
if(Data.SAMTOOLS() && useStreamer){ReadWrite.USE_SAMBAMBA=false;} //Disable because it takes forever to read the header
}
ByteFile tf=ByteFile.makeByteFile(in1, false);
// System.err.println("B");
final ByteStreamWriter tsw=(outsam==null ? null : new ByteStreamWriter(outsam, overwrite, false, true));
if(tsw!=null){tsw.start();}
ReadWrite.USE_SAMBAMBA=true;
// System.err.println("C");
processHeader(tf, tsw);
// System.err.println("D");
processReference();
// System.err.println("E");
if(maxReads<0){maxReads=Long.MAX_VALUE;}
if(covIn!=null && !covIn.isEmpty()) {
for(String s : covIn) {
loadCoverageFile(s);
}
}else if(useStreamer && !FileFormat.isStdio(in1)){
tf.close();
// System.err.println("F");
processViaStreamer(tsw);
// System.err.println("G");
}else{
// processViaByteFile(tf, line, tsw);
processViaByteFile(tf, tsw);
// System.err.println("H");
}
// System.err.println("I");
printOutput();
if(orffasta!=null){
processOrfsFasta(orffasta, outorf, table);
}
if(tsw!=null){tsw.waitForFinish();}
}
private void processViaStreamer(ByteStreamWriter tsw){
SamLineStreamer ss=new SamLineStreamer(in1, streamerThreads, false, maxReads);
ss.start();
ByteBuilder bb=new ByteBuilder(33000);
for(ListNum<SamLine> ln=ss.nextLines(); ln!=null && ln.size()>0; ln=ss.nextLines()){
ArrayList<SamLine> list=(ln==null ? null : ln.list);
for(SamLine sl : list){
if(tsw!=null){
sl.toBytes(bb);
bb.nl();
if(bb.length>=16384){
tsw.print(bb);
bb.clear();
}
}
processSamLine(sl);
}
}
if(tsw!=null){
if(bb.length>0){tsw.print(bb);}
tsw.poison();
}
}
private void processViaByteFile(ByteFile tf, ByteStreamWriter tsw){
for(byte[] line=tf.nextLine(); line!=null && readsProcessed<maxReads; line=tf.nextLine()){
if(tsw!=null){tsw.println(line);}
processSamLine(line);
}
tf.close();
if(tsw!=null){tsw.poison();}
}
//Untested
//See also CoverageArray.loadDepth; that implementation is better (also untested)
private void loadCoverageFile(String fname) {
ByteFile bf=ByteFile.makeByteFile(fname, true);
byte[] line=bf.nextLine();
for(; line!=null && Tools.startsWith(line, '#'); line=bf.nextLine()) {} //Skip header
if(line==null) {return;}
lp.set(line);
String name=lp.parseString(0);
Scaffold scaf=table.get(name);
for(; line!=null; line=bf.nextLine()) {
assert(!Tools.startsWith(line, '#'));
lp.set(line);
int len=lp.length(0);
if(len>0 && !lp.termEquals(name, 0)) {
name=lp.parseString(0);
scaf=table.get(name);
}
int pos=lp.parseInt(1);
int cov=lp.parseInt(2);
if(cov>0) {
if(USE_COVERAGE_ARRAYS){
if(scaf.obj0==null){
scaf.obj0=(bits32 ? new CoverageArray3(table.size(), scaf.length) : new CoverageArray2(table.size(), scaf.length));
}
CoverageArray ca=(CoverageArray)scaf.obj0;
ca.increment(pos, cov);
}else if(USE_BITSETS){
if(scaf.obj0==null){scaf.obj0=new BitSet(scaf.length+1);}
BitSet bs=(BitSet)scaf.obj0;
bs.set(pos);
}
}
}
}
/*--------------------------------------------------------------*/
/*---------------- Setup ----------------*/
/*--------------------------------------------------------------*/
/** Process all sam header lines from the tf.
* Once a non-header line is encountered, return it.
* If non-null, print all lines to the tsw. */
public void processHeader(ByteFile tf, ByteStreamWriter tsw){
byte[] line=null;
for(line=tf.nextLine(); line!=null && (line.length==0 || line[0]=='@'); line=tf.nextLine()){
// System.err.println(new String(line));
if(tsw!=null){tsw.println(line);}
if(line.length>2){
final byte a=line[1], b=line[2];
if(a=='S' && b=='Q'){
lp.set(line);
Scaffold scaf=new Scaffold(lp);
if(COUNT_GC){scaf.basecount=KillSwitch.allocLong1D(8);}
assert(!table.containsKey(scaf.name)) : "\nDuplicate scaffold name!\n"+scaf+"\n\n"+table.get(scaf.name);
table.put(scaf.name, scaf);
list.add(scaf);
refBases+=scaf.length;
// sc.obj=new CoverageArray2(table.size(), sc.length+1);
// outstream.println("Made scaffold "+sc.name+" of length "+sc.length);
}else if(a=='P' && b=='G'){
lp.set(line);
for(int i=1, terms=lp.terms(); i<terms; i++) {
if(lp.termStartsWith("PN:", i)){
if(program==null){
lp.incrementA(3);
program=lp.parseStringFromCurrentField();
}
}else if(lp.termStartsWith("VN:", i)){
if(version==null){
lp.incrementA(3);
version=lp.parseStringFromCurrentField();
}
}
}
}else if(a=='R' && b=='G'){
//Do nothing
}else if(a=='H' && b=='D'){
//Do nothing
}else if(a=='C' && b=='O'){
//Do nothing
}else{
// assert(false) : line;
}
}
}
if(line!=null){tf.pushBack(line);}
// return line;
}
public void loadScaffoldsFromIndex(int minChrom, int maxChrom){
final int[][] lengths=Data.scaffoldLengths;
final int[][] locs=Data.scaffoldLocs;
final byte[][][] names=Data.scaffoldNames;
final int[] counts=new int[8];
for(int chrom=minChrom; chrom<=maxChrom; chrom++){
final ChromosomeArray ca=Data.getChromosome(chrom);
// assert(false) : lengths[chrom]+", "+lengths.length+", "+names[chrom]+", "+locs[chrom]+", "+(ca==null);
if(lengths[chrom]!=null){
final int[] clengths=lengths[chrom];
final int[] clocs=locs[chrom];
final byte[][] cnames=names[chrom];
for(int idx=0; idx<clengths.length; idx++){
final int length=clengths[idx];
final int loc=clocs[idx];
final String name=new String(cnames[idx]);
final Scaffold scaf=new Scaffold(name, length);
if(ca!=null){
scaf.gc=ca.calcGC(loc, length, counts);
}
if(COUNT_GC){scaf.basecount=KillSwitch.allocLong1D(8);}
assert(!table.containsKey(scaf.name)) : "\nDuplicate scaffold name!\n"+scaf+"\n\n"+table.get(scaf.name);
table.put(scaf.name, scaf);
list.add(scaf);
refBases+=scaf.length;
}
}
}
}
public void processReference(){
if(reference==null){return;}
ByteFile bf=ByteFile.makeByteFile(reference, false);
Scaffold scaf=null;
int len=0;
final long[] acgtn=KillSwitch.allocLong1D(8);
boolean addLen=false;
for(byte[] s=bf.nextLine(); s!=null; s=bf.nextLine()){
if(s.length>0 && s[0]=='>'){
if(scaf!=null){
if(scaf.length>0 && scaf.length!=len){
outstream.println("ERROR: Scaffold "+scaf.name+" has contradictory lengths of "+scaf.length+" and "+len+"\n"
+ "This probably indicates a corrupt or incorrect reference.");
errorState=true;
if(Shared.EA()){KillSwitch.kill();}
}
// else{outstream.println(scaf.name+", "+scaf.length+", "+len);}
scaf.length=len;
if(addLen){
refBases+=scaf.length;
addLen=false;
}
scaf.gc=(float)((acgtn[1]+acgtn[2])*1d/Data.max(1, acgtn[0]+acgtn[1]+acgtn[2]+acgtn[3]));
scaf=null;
len=0;
Arrays.fill(acgtn, 0);
}
String name=new String(s, 1, s.length-1);
scaf=table.get(name);
if(ADD_FROM_REF && scaf==null){
scaf=new Scaffold(name, 0);
if(!warned){
outstream.println("Warning - SAM header did not include "+name+"\nAbsent scaffolds will be added; further warnings will be suppressed.");
warned=true;
}
if(COUNT_GC){scaf.basecount=KillSwitch.allocLong1D(8);}
table.put(name, scaf);
list.add(scaf);
addLen=true;
}
}else{
len+=s.length;
for(int i=0; i<s.length; i++){
acgtn[charToNum[s[i]]]++;
}
}
}
if(scaf!=null){
if(scaf.length>0 && scaf.length!=len){
outstream.println("ERROR: Scaffold "+scaf.name+" has contradictory lengths of "+scaf.length+" and "+len+"\n"
+ "This probably indicates a corrupt or incorrect reference.");
errorState=true;
if(Shared.EA()){KillSwitch.kill();}
}
// else{outstream.println(scaf.name+", "+scaf.length+", "+len);}
scaf.length=len;
if(addLen){
refBases+=scaf.length;
addLen=false;
}
scaf.gc=(float)((acgtn[1]+acgtn[2])*1d/Data.max(1, acgtn[0]+acgtn[1]+acgtn[2]+acgtn[3]));
scaf=null;
len=0;
Arrays.fill(acgtn, 0);
}
}
public void processOrfsFasta(String fname_in, String fname_out, HashMap<String, Scaffold> map){
TextFile tf=new TextFile(fname_in, false);
assert(!fname_in.equalsIgnoreCase(fname_out));
TextStreamWriter tsw=new TextStreamWriter(fname_out, overwrite, false, true);
tsw.start();
if(printHeader){
String pound=(headerPound ? "#" : "");
tsw.print(pound+"mappedBases="+mappedBases+"\n");
tsw.print(pound+"mappedNonClippedBases="+mappedNonClippedBases+"\n");
tsw.print(pound+"mappedBasesWithDels="+mappedBasesWithDels+"\n");
tsw.print(pound+"mappedReads="+mappedReads+"\n");
tsw.print(pound+"name\tlength\tdepthSum\tavgDepth\tavgDepth/mappedBases\tminDepth\tmaxDepth\tmedianDepth\tstdDevDepth\tfractionCovered\n");
}
String line;
final StringBuilder sb=new StringBuilder(500);
// Formatter formatter=new Formatter(sb);
while((line=tf.nextLine())!=null){
if(line.length()>1 && line.charAt(0)=='>'){
String[] split=line.split(" # "); //' # ' used as delimiters
String orfname=split[0].substring(1).trim(); //In case there are spaces around the ' # ' delimiters
String scafname=orfname;
if(scafname.contains("_")){//PRODIGAL pads _1 to the name of the first orf of a scaffold, and etc
int last=scafname.lastIndexOf('_');
boolean numeric=false;
for(int i=last+1; i<scafname.length(); i++){
if(Tools.isDigit(scafname.charAt(i))){numeric=true;}
else{numeric=false; break;}
}
if(numeric){scafname=scafname.substring(0, last);}
}
int start=Integer.parseInt(split[1].trim());
int stop=Integer.parseInt(split[2].trim());
int strand=Integer.parseInt(split[3].trim());
if(strand==1){strand=Shared.PLUS;}else{strand=Shared.MINUS;}
Orf orf=new Orf(orfname, start, stop, (byte)strand);
Scaffold scaf=map.get(scafname);
// if(scaf==null){scaf=map.get(orfname);}
// assert(scaf!=null) : "\nCan't find scaffold for ("+orf+")\nfrom line\n"+line+"\n";
// assert(orf.start>=0 && orf.stop<scaf.length) : "\norf goes out of scaffold bounds.\n"+orf+"\n"+scaf+"\n";
if(scaf==null){
outstream.println("Can't find scaffold for ("+orf+")\nfrom line\n"+line+"\nscafname='"+scafname+"'\norfname='"+orfname+"'");
if(ABORT_ON_ERROR){
tsw.poison();
throw new RuntimeException("Aborting.");
}
}else{
if(orf.start<0 && orf.stop>=scaf.length){
outstream.println("orf goes out of scaffold bounds.\n"+orf+"\n"+scaf);
if(ABORT_ON_ERROR){
tsw.poison();
throw new RuntimeException("Aborting.");
}
}
CoverageArray ca=(CoverageArray)(STRANDED && strand==1 ? scaf.obj1 : scaf.obj0); //TODO: Strand logic here depends on stranding protocol.
orf.readCoverageArray(ca);
}
// {
// tsw.print(Tools.format("%s\t%d\t", args));
// }
sb.append(orf.name).append('\t');
sb.append(orf.length()).append('\t');
sb.append(orf.baseDepth).append('\t');
sb.append(Tools.format("%.4f", orf.avgCoverage())).append('\t');
sb.append(orf.avgCoverage()/mappedNonClippedBases);
sb.append('\t');
sb.append(orf.minDepth).append('\t');
sb.append(orf.maxDepth).append('\t');
sb.append(orf.medianDepth).append('\t');
sb.append(Tools.format("%.4f",orf.stdevDepth)).append('\t');
sb.append(Tools.format("%.4f",orf.fractionCovered()));
sb.append('\n');
tsw.print(sb.toString());
sb.setLength(0);
}
}
tsw.poisonAndWait();
}
/*--------------------------------------------------------------*/
/*---------------- Inner Methods ----------------*/
/*--------------------------------------------------------------*/
public boolean addCoverage(final String scafName, final byte[] seq, byte[] match, final int start0, final int stop0, final int readlen,
final int nonClippedBases, final int strand, int incrementFrags, boolean properPair, SamLine sl){//sl is optional
Scaffold scaf=table.get(scafName);
if(scaf==null){
if(ADD_FROM_READS){
if(!warned){
outstream.println("Warning: A read was mapped to unknown reference sequence "+scafName+
"\nFurther warnings will be suppressed. These scaffolds will be included, but \n"
+ "some coverage statistics will be inaccurate as scaffold lengths are not known.");
warned=true;
}
scaf=new Scaffold(scafName, 0);
if(COUNT_GC){scaf.basecount=KillSwitch.allocLong1D(8);}
table.put(scafName, scaf);
list.add(scaf);
return addCoverage(scaf, seq, match, start0, stop0, readlen, nonClippedBases, strand, incrementFrags, properPair, sl);
}else if(EA){
KillSwitch.kill("ERROR: A read was mapped to unknown reference sequence "+scafName);
}else if(!warned){
outstream.println("Warning: Can't find "+scafName+"\nThis scaffold will not be included. Further warnings will be suppressed.");
warned=true;
error=true;
}
return false;
}
return addCoverage(scaf, seq, match, start0, stop0, readlen, nonClippedBases, strand, incrementFrags, properPair, sl);
}
public boolean addCoverage(final Scaffold scaf, final byte[] seq, byte match[], final int start0, final int stop0, final int readlen, final int nonClippedBases,
final int strand, int incrementFrags, boolean properPair, SamLine sl){//sl is optional
if(scaf==null){
assert(false) : "Adding coverage to a null Scaffold.";
return false;
}
if(ADD_FROM_READS){
int x=scaf.length;
scaf.length=Tools.max(scaf.length, stop0+1);
if(scaf.length!=x){
refBases=refBases+scaf.length-x;
}
}
final int start=Tools.max(start0, 0);
final int stop=Tools.min(stop0, scaf.length-1);
assert(start>=0 && stop>=0) : "\nAn error was encountered when processing a read. Output will not be valid.\n"+
"\nscafName="+scaf.name+"\nseq="+new String(seq)+"\nstart="+start+
"\nstop="+stop+"\nreadlen="+readlen+"\nstrand="+strand+"\nscaf.length="+scaf.length+"\nscaf="+scaf;
mappedBases+=readlen;
mappedNonClippedBases+=nonClippedBases;
mappedBasesWithDels+=(stop-start+1);
mappedReads++;
if(properPair){properPairs++;}
scaf.readhits++;
scaf.fraghits+=incrementFrags;
if(strand==1){scaf.readhitsMinus++;}
if(seq!=null && scaf.basecount!=null){
final long[] counts=scaf.basecount;
for(int i=0; i<seq.length; i++){
counts[charToNum[seq[i]]]++;
}
}
if(!INCLUDE_DELETIONS && !START_ONLY && !STOP_ONLY){
assert(match!=null) : "Coverage excluding deletions cannot be calculated without a match string.";
return addCoverageIgnoringDeletions(scaf, seq, match, start, stop, readlen, strand, incrementFrags);
}
final int basehits=stop-start+1;
scaf.basehits+=basehits;
if(USE_COVERAGE_ARRAYS){
if(scaf.obj0==null){
scaf.obj0=(bits32 ? new CoverageArray3(table.size(), scaf.length) : new CoverageArray2(table.size(), scaf.length));
if(STRANDED){
scaf.obj1=(bits32 ? new CoverageArray3(table.size(), scaf.length) : new CoverageArray2(table.size(), scaf.length));
}
}
CoverageArray ca=(CoverageArray)(STRANDED && strand==1 ? scaf.obj1 : scaf.obj0);
if(START_ONLY){
ca.increment(start);
}else if(STOP_ONLY){
ca.increment(stop);
}else{
ca.incrementRange(start, stop);
// mappedBases+=(stop-start+1);
// mappedBases+=(readlen);
// assert(readlen==(stop-start+1)) : "readlen="+readlen+", (stop-start+1)="+(stop-start+1)+", start="+start+", stop="+stop+", start0="+start0+", stop0="+stop0+
// "\n"+sl+"\n";
//123
}
}else if(USE_BITSETS){
if(scaf.obj0==null){
scaf.obj0=new BitSet(scaf.length+1);
if(STRANDED){
scaf.obj1=new BitSet(scaf.length+1);
}
}
BitSet bs=(BitSet)(STRANDED && strand==1 ? scaf.obj1 : scaf.obj0);
if(START_ONLY){
bs.set(start);
}else if(STOP_ONLY){
bs.set(stop);
}else{
bs.set(start, stop+1);
}
}
return true;
}
private boolean addCoverageIgnoringDeletions(final Scaffold scaf, final byte[] seq, byte match[], final int start, final int stop, final int readlen, final int strand, int incrementFrags){
assert(!INCLUDE_DELETIONS && !START_ONLY && !STOP_ONLY);
assert(match!=null) : "Coverage excluding deletions cannot be calculated without a match string.";
if(Read.isShortMatchString(match)){
match=Read.toLongMatchString(match);
}
int basehits=0;
if(USE_COVERAGE_ARRAYS){
if(scaf.obj0==null){
scaf.obj0=(bits32 ? new CoverageArray3(table.size(), scaf.length) : new CoverageArray2(table.size(), scaf.length));
if(STRANDED){
scaf.obj1=(bits32 ? new CoverageArray3(table.size(), scaf.length) : new CoverageArray2(table.size(), scaf.length));
}
}
CoverageArray ca=(CoverageArray)(STRANDED && strand==1 ? scaf.obj1 : scaf.obj0);
for(int rpos=start, mpos=0; mpos<match.length && rpos<=stop; mpos++){
byte m=match[mpos];
if(m=='m' || m=='S' || m=='N'){
ca.increment(rpos, 1);
basehits++;
rpos++;
}else if(m=='X' || m=='Y' || m=='C' || m=='I'){
//do nothing
}else if(m=='D'){
rpos++;
}else{
assert(false) : "Unhandled symbol "+m;
}
}
}else if(USE_BITSETS){
if(scaf.obj0==null){
scaf.obj0=new BitSet(scaf.length+1);
if(STRANDED){
scaf.obj1=new BitSet(scaf.length+1);
}
}
BitSet bs=(BitSet)(STRANDED && strand==1 ? scaf.obj1 : scaf.obj0);
for(int rpos=start, mpos=0; mpos<match.length && rpos<=stop; mpos++){
byte m=match[mpos];
if(m=='m' || m=='S' || m=='N'){
bs.set(rpos);
basehits++;
rpos++;
}else if(m=='X' || m=='Y' || m=='C' || m=='I'){
//do nothing
}else if(m=='D'){
rpos++;
}else{
assert(false) : "Unhandled symbol "+m;
}
}
}
scaf.basehits+=basehits;
return true;
}
public boolean processSamLine(byte[] line){
if(line==null || line.length<3){
return false;
}else if(line[0]=='@'){
if(!error){
outstream.println("Unexpected header line: "+line);
outstream.println("This should not cause problems, and is probably due to concatenated sam files.\n" +
"Supressing future unexpected header warnings.");
error=true;
}
if(line[1]=='S' && line[2]=='Q'){
lp.set(line);
String name=Scaffold.name(lp);
if(!table.containsKey(name)){
Scaffold scaf=new Scaffold(lp);
if(COUNT_GC){scaf.basecount=KillSwitch.allocLong1D(8);}
table.put(scaf.name, scaf);
list.add(scaf);
refBases+=scaf.length;
}
}
}else{
SamLine sl=new SamLine(line);
return processSamLine(sl);
}
return false;
}
private int trim(SamLine sl){
assert(border>0 || (trimq>=0 && (qtrimLeft || qtrimRight)));
Read r=null;
Scaffold sc=null;
int leftTrimAmount=border, rightTrimAmount=border;
if(border>0){
r=sl.toRead(false);
sc=table.get(sl.rnameS());
assert(sc!=null) : sl+"\n\n"+sl.rnameS()+"\n\n"+table;
int skipTrimRange=Tools.max(10, border+5);
if(r.start<skipTrimRange){
if(r.strand()==Shared.PLUS){leftTrimAmount=0;}
else{rightTrimAmount=0;}
}
if(r.stop>sc.length-skipTrimRange){
if(r.strand()==Shared.PLUS){rightTrimAmount=0;}
else{leftTrimAmount=0;}
}
}
final int len0=sl.length();
if(qtrimLeft || qtrimRight){
long packed=TrimRead.testOptimal(sl.seq, sl.qual, trimE);
if(qtrimLeft){leftTrimAmount=Tools.max(leftTrimAmount, (int)((packed>>32)&0xFFFFFFFFL));}
if(qtrimRight){rightTrimAmount=Tools.max(rightTrimAmount, (int)((packed)&0xFFFFFFFFL));}
// assert(false) : qtrimLeft+", "+qtrimRight+", "+trimq+", "+trimE+", "+rightTrimAmount+"\n"+sl+"\n";
}
final int trimmed;
if(leftTrimAmount<1 && rightTrimAmount<1){trimmed=0;}
else{
if(r==null){r=sl.toRead(false);}
if(sc==null){sc=table.get(sl.rnameS());}
int scaflen=(sc==null ? 1999999999 : sc.length);
trimmed=TrimRead.trimReadWithMatch(r, sl, leftTrimAmount, rightTrimAmount, 0, scaflen, false);
}
// assert(trimmed==len0-sl.length()) : trimmed+", "+len0+", "+sl.length();
// assert(rightTrimAmount>0) : qtrimLeft+", "+qtrimRight+", "+trimq+", "+trimE+", "+rightTrimAmount+"\n"+sl+"\n";
return trimmed;
}
public boolean processSamLine(SamLine sl){
readsProcessed++;
basesProcessed+=sl.length();
if(sl.duplicate() && !INCLUDE_DUPLICATES){return false;}
if(border>0 || (trimq>=0 && (qtrimLeft || qtrimRight))){
if(sl.mapped() && sl.seq!=null){
int trimmed=trim(sl);
if(sl.length()<1 || trimmed<0){return false;}//trimmed<0 implies everything was trimmed
}
}
final int kmers=Tools.countKmers(sl.seq, k);
final double cKmers=sl.qual==null ? kmers : Tools.countCorrectKmers(sl.qual, k);
kmersProcessed+=kmers;
correctKmers+=cKmers;
final boolean properPair=(sl.hasMate() && sl.mapped() && sl.primary() && sl.properPair());
if(PHYSICAL_COVERAGE && properPair){
SamLine mate=pairTable.remove(sl.qname);
if(mate==null){pairTable.put(sl.qname, sl);}
else{
final int start1=sl.start(INCLUDE_SOFT_CLIP, false);
final int stop1=sl.stop(start1, INCLUDE_SOFT_CLIP, false);
final int start2=mate.start(INCLUDE_SOFT_CLIP, false);
final int stop2=mate.stop(start2, INCLUDE_SOFT_CLIP, false);
final int strand=(sl.pairnum()==0 ? sl.strand() : mate.strand());
final int length=USE_TLEN ? sl.tlen : Tools.max(stop1, stop2)-Tools.min(start1, start2)+1;
mappedKmers+=kmers;
addCoverage(sl.rnameS(), null, null, Tools.min(start1, start2), Tools.max(stop1, stop2), length, sl.mappedNonClippedBases(), strand, 2, sl.properPair(), sl);
}
}else if(sl.mapped() && (USE_SECONDARY || sl.primary()) && sl.mapq>=minMapq){
assert(sl.seq!=null || sl.cigar!=null) : "This program requires bases or a cigar string for every sam line. Problem line:\n"+sl+"\n";
// assert(sl.seq!=null) : sl.toString();
final int length=sl.length();
final int start=sl.start(INCLUDE_SOFT_CLIP, false);
final int stop=sl.stop(start, INCLUDE_SOFT_CLIP, false);
// assert(false && length==stop-start+1) : length+", "+start+", "+stop+", "+(stop-start+1);
// assert(false) : "'"+new String(sl.rname())+"', '"+sl.rnameS()+"'";
// assert(false) : "'"+sl.rnameS()+"'";
final byte[] match=(INCLUDE_DELETIONS ? null : sl.toShortMatch(true));
mappedKmers+=kmers;
return addCoverage(sl.rnameS(), sl.seq, match, start, stop, length, sl.mappedNonClippedBases(), sl.strand(), sl.hasMate() ? 1 : 2, sl.properPair(), sl);
}
return false;
}
public boolean processRead(Read r){
readsProcessed++;
final int kmers=Tools.countKmers(r.bases, k);
final double cKmers=r.quality==null ? kmers : Tools.countCorrectKmers(r.quality, k);
kmersProcessed+=kmers;
correctKmers+=cKmers;
if(r.mapped() && r.bases!=null){
if(USE_SECONDARY && r.sites!=null && r.sites.size()>0){
boolean b=false;
for(SiteScore ss : r.sites){
b=processRead(r, ss) || b;
}
return b;
}else{
final Read mate=r.mate;
final boolean set1=coords.set(r);
final boolean set2=(PHYSICAL_COVERAGE && r.paired() && r.pairnum()==0 && coords2.set(mate));
if(set1 && set2 && Tools.equals(coords.name, coords2.name)){
final int start1=coords.start;
final int stop1=coords.stop;
final int start2=coords2.start;
final int stop2=coords2.stop;
final int strand=r.strand();
final int length=Tools.max(stop1, stop2)-Tools.min(start1, start2)+1;
mappedKmers+=kmers;
addCoverage(new String(coords.name), null, null, Tools.min(start1, start2), Tools.max(stop1, stop2), length,
r.mappedNonClippedBases(), strand, 2-r.mateCount(), r.paired(), null);
}else{
if(set1){
mappedKmers+=kmers;
return addCoverage(new String(coords.name), r.bases, r.match, coords.start, coords.stop, r.length(),
r.mappedNonClippedBases(), coords.strand, 2-r.mateCount(), r.paired(), null);
}
}
}
}
return false;
}
public boolean processRead(Read r, SiteScore ss){
if(ss!=null && r.bases!=null){
if(coords.set(ss)){
return addCoverage(new String(coords.name), r.bases, ss.match, coords.start, coords.stop, r.length(),
r.mappedNonClippedBases(), coords.strand, 1, r.paired(), null);
}
}
return false;
}
/*--------------------------------------------------------------*/
/*---------------- Output Methods ----------------*/
/*--------------------------------------------------------------*/
public void printOutput(){
totalScaffolds=list.size();
long refKmers=0;
long refBases2=0;
for(Scaffold sc : list){
refBases2+=sc.length;
if(sc.length>=k){refKmers+=sc.length-k+1;}
}
String basecov1=(basecov==null ? null : (STRANDED ? basecov.replaceFirst("#", "1") : basecov));
String rangecov1=(rangecov==null ? null : (STRANDED ? rangecov.replaceFirst("#", "1") : rangecov));
String bincov1=(bincov==null ? null : (STRANDED ? bincov.replaceFirst("#", "1") : bincov));
String normcov1=(normcov==null ? null : (STRANDED ? normcov.replaceFirst("#", "1") : normcov));
String normcovOverall1=(normcovOverall==null ? null : (STRANDED ? normcovOverall.replaceFirst("#", "1") : normcovOverall));
String histogram1=(histogram==null ? null : (STRANDED ? histogram.replaceFirst("#", "1") : histogram));
String stats1=(covstats==null ? null : (STRANDED ? covstats.replaceFirst("#", "1") : covstats));
String basecov2=(basecov==null || !STRANDED ? null : basecov.replaceFirst("#", "2"));
String rangecov2=(rangecov==null || !STRANDED ? null : rangecov.replaceFirst("#", "2"));
String bincov2=(bincov==null || !STRANDED ? null : bincov.replaceFirst("#", "2"));
String normcov2=(normcov==null || !STRANDED ? null : normcov.replaceFirst("#", "2"));
String normcovOverall2=(normcovOverall==null ? null : (STRANDED ? normcovOverall.replaceFirst("#", "2") : normcovOverall));
String histogram2=(histogram==null || !STRANDED ? null : histogram.replaceFirst("#", "2"));
String stats2=(covstats==null || !STRANDED ? null : covstats.replaceFirst("#", "2"));
if(CONCISE){
writeCoveragePerBaseConcise(basecov1, list, 0, minscaf);
writeCoveragePerBaseConcise(basecov2, list, 1, minscaf);
}else{
writeCoveragePerBase(basecov1, list, DELTA_ONLY, 0, minscaf);
writeCoveragePerBase(basecov2, list, DELTA_ONLY, 1, minscaf);
}
if(KEEP_SHORT_BINS){
writeCoveragePerBaseBinned2(bincov1, list, binsize, 0, minscaf);
writeCoveragePerBaseBinned2(bincov2, list, binsize, 1, minscaf);
}else{
writeCoveragePerBaseBinned(bincov1, list, binsize, 0, minscaf);
writeCoveragePerBaseBinned(bincov2, list, binsize, 1, minscaf);
}
if(rangecov!=null) {
writeCoverageRanges(rangecov1, list, 0, minscaf);
writeCoverageRanges(rangecov2, list, 1, minscaf);
}
if(normcov!=null){
writeCoveragePerBaseNormalized(normcov1, list, binsize, 0, minscaf);
writeCoveragePerBaseNormalized(normcov2, list, binsize, 1, minscaf);
}
if(normcovOverall!=null){
writeCoveragePerBaseNormalizedOverall(normcovOverall1, list, binsize, 0, minscaf);
writeCoveragePerBaseNormalizedOverall(normcovOverall2, list, binsize, 1, minscaf);
}
if(outrpkm!=null){
writeRPKM(outrpkm, in1, null, readsProcessed, NONZERO_ONLY,list);
}
{
long[] hist=writeStats(stats1, 0);
if(hist!=null){writeHist(histogram1, hist);}
if(STRANDED){
hist=writeStats(stats2, 1);
if(hist!=null){writeHist(histogram2, hist);}
}
}
final double mult=1.0/refBases;
double depthCovered=mappedBases*mult;
double depthCovered2=mappedNonClippedBases*mult;
double depthCovered3=mappedBasesWithDels*mult;
double pctScaffoldsWithCoverage=scaffoldsWithCoverage1*100.0/totalScaffolds;
double pctCovered=totalCoveredBases1*100*mult;
if(!KEY_VALUE){
outstream.println("Reads: \t"+readsProcessed);
outstream.println("Mapped reads: \t"+mappedReads);
// outstream.println("Mapped bases: \t"+mappedBases);
// outstream.println("Mapped non-clipped bases: \t"+mappedNonClippedBases);
outstream.println("Mapped bases: \t"+mappedNonClippedBases);
outstream.println("Ref scaffolds: \t"+totalScaffolds);
outstream.println("Ref bases: \t"+refBases);
if(k>0){
String kcovS=k+"-mer coverage:";
String kcorrectS="Percent correct "+k+"-mers:";
while(kcovS.length()<26){kcovS=kcovS+" ";}
while(kcovS.length()<26){kcorrectS=kcorrectS+" ";}
outstream.println(Tools.format("\n"+kcovS+" \t%.3f", mappedKmers*1.0/refKmers));
outstream.println(Tools.format(kcorrectS+" \t%.3f", 100*correctKmers/kmersProcessed));
// outstream.println(kmersProcessed+", "+correctKmers);
}
outstream.println(Tools.format("\nPercent mapped: \t%.3f", mappedReads*100f/readsProcessed));
outstream.println(Tools.format("Percent proper pairs: \t%.3f", properPairs*100f/readsProcessed));
// outstream.println(depthCovered);
outstream.println(Tools.format("Average coverage: \t%.3f", depthCovered2));
outstream.println(Tools.format("Average coverage with deletions: \t%.3f", depthCovered3));
if(USE_COVERAGE_ARRAYS && calcCovStdev){
double[] stdev=standardDeviation(list, 0, minscaf);
outstream.println(Tools.format("Standard deviation: \t%.3f", stdev[1]));
}
outstream.println(Tools.format("Percent scaffolds with any coverage: \t%.2f", pctScaffoldsWithCoverage));
if(USE_COVERAGE_ARRAYS || USE_BITSETS){
outstream.println(Tools.format("Percent of reference bases covered: \t%.2f", pctCovered));
}
}else{
outstream.println("reads="+readsProcessed);
outstream.println("mappedReads="+mappedReads);
outstream.println("mappedBases="+mappedNonClippedBases);
outstream.println("mappedBasesWithDels="+mappedBasesWithDels);
outstream.println("refScaffolds="+totalScaffolds);
outstream.println("refBases="+refBases);
outstream.println(Tools.format("percentMapped=%.3f", mappedReads*100f/readsProcessed));
outstream.println(Tools.format("percentPaired=%.3f", properPairs*100f/readsProcessed));
outstream.println(Tools.format("averageCoverage=%.3f", depthCovered2));
if(USE_COVERAGE_ARRAYS && calcCovStdev){
double[] stdev=standardDeviation(list, 0, minscaf);
outstream.println(Tools.format("standardDeviation=%.3f", stdev[1]));
}
outstream.println(Tools.format("percentCoveredScaffolds=%.2f", pctScaffoldsWithCoverage));
if(USE_COVERAGE_ARRAYS || USE_BITSETS){
outstream.println(Tools.format("percentCoveredBases=%.2f", pctCovered));
}
}
}
public int basesUnderAverageCoverage(final int[] array, final double avg, final int window){
if(array.length<window){return 0;}
final long limit=(long)Math.ceil(window*avg);
long covSum=0;
int baseCount=0;
for(int i=0; i<window; i++){
covSum+=array[i];
}
boolean below=false;
int lastStop=-1, lastStart=0;
for(int a=0, b=window; b<array.length; a++, b++){
if(covSum>=limit){
if(below){//end range
baseCount=b-Tools.max(lastStop+1, lastStart);
lastStop=b-1;
below=false;
}
}else{
if(!below){//start range
lastStart=a;
below=true;
}
}
covSum-=array[a];
assert(covSum>=0);
covSum+=array[b];
}
if(below){//end range
baseCount=array.length-Tools.max(lastStop, lastStart);
}
assert(baseCount>=0);
return baseCount;
}
public int basesUnderAverageCoverage(final char[] array, final double avg, final int window){
if(array.length<window){return 0;}
final long limit=(long)Math.ceil(window*avg);
long covSum=0;
int baseCount=0;
for(int i=0; i<window; i++){
covSum+=array[i];
}
// outstream.println("limit: "+limit);
boolean below=false;
int lastStop=-1, lastStart=0;
for(int a=0, b=window; b<array.length; a++, b++){
if(covSum>=limit){
if(below){//end range
baseCount+=b-Tools.max(lastStop+1, lastStart);
// outstream.println("\nprev: "+lastStop+", "+lastStart);
// outstream.println("end range at "+a+", "+b);
// outstream.println("baseCount: "+baseCount+", covSum="+covSum);
lastStop=b-1;
below=false;
}
}else{
if(!below){//start range
// outstream.println("\nprev: "+lastStop+", "+lastStart);
// outstream.println("start range at "+a+", "+b);
// outstream.println("baseCount: "+baseCount+", covSum="+covSum);
lastStart=a;
below=true;
}
}
covSum-=array[a];
assert(covSum>=0);
covSum+=array[b];
}
if(below){//end range
baseCount+=array.length-Tools.max(lastStop+1, lastStart);
// outstream.println("\nprev: "+lastStop+", "+lastStart);
// outstream.println("end range at "+array.length);
// outstream.println("baseCount: "+baseCount+", covSum="+covSum);
}
assert(baseCount>=0);
return baseCount;
}
public long[] writeStats(String fname, int strand){
// outstream.println("Writing stats for "+fname+", "+strand);
final TextStreamWriter tsw=(fname==null ? null : new TextStreamWriter(fname, overwrite, false, true));
if(tsw!=null){
tsw.start();
if(printHeader){
String pound=(headerPound ? "#" : "");
if(TWOCOLUMN){
tsw.println(pound+"ID\tAvg_fold");
}else{
tsw.println(pound+"ID\tAvg_fold\tLength\tRef_GC\tCovered_percent\tCovered_bases\tPlus_reads\tMinus_reads"+
(COUNT_GC ? "\tRead_GC" : "")+
(USE_COVERAGE_ARRAYS ? ("\tMedian_fold\tStd_Dev") : "")+
(USE_WINDOW ? "\tUnder_"+Tools.format("%.0f",LOW_COV_DEPTH)+"/"+LOW_COV_WINDOW : ""));
}
}
//Maximally:
//"ID\tAvg_fold\tLength\tRef_GC\tCovered_percent\tCovered_bases\tPlus_reads\tMinus_reads\tRead_GC\tMedian_fold\tStd_Dev\tUnder_X"
}
final int histmax=(bits32 ? 1000000 : Character.MAX_VALUE);
final LongList hist=new LongList(Character.MAX_VALUE);
long coveredScafTemp=0;
long coveredBaseTemp=0;
for(Scaffold scaf : list){
final long sum=scaf.basehits;
int covered=0;
int median=0;
int underWindowAverage=0;
final double stdev;
if(USE_COVERAGE_ARRAYS){
CoverageArray ca=(CoverageArray)(STRANDED && strand==1 ? scaf.obj1 : scaf.obj0);
if(ca!=null){
for(int i=0; i<scaf.length; i++){
int x=ca.get(i);
hist.increment(Tools.min(x, histmax));
// sum+=x;
if(x>=minDepthToBeCovered){covered++;}
}
if(bits32){
int[] array=((CoverageArray3)ca).array;
stdev=Tools.standardDeviation(array);
underWindowAverage=basesUnderAverageCoverage(array, LOW_COV_DEPTH, LOW_COV_WINDOW);
Shared.sort(array);
Tools.reverseInPlace(array);
median=ca.get(scaf.length/2);
}else{
char[] array=((CoverageArray2)ca).array;
stdev=Tools.standardDeviation(array);
underWindowAverage=basesUnderAverageCoverage(array, LOW_COV_DEPTH, LOW_COV_WINDOW);
Arrays.sort(array);
Tools.reverseInPlace(array);
median=ca.get(scaf.length/2);
}
}else{
hist.increment(0, scaf.length);
stdev=0;
}
}else if(USE_BITSETS){
// sum+=scaf.basehits;
BitSet bs=(BitSet)(STRANDED && strand==1 ? scaf.obj1 : scaf.obj0);
covered=(bs==null ? 0 : bs.cardinality());
stdev=-1;
}else{
stdev=-1;
}
if(sum>0){
coveredScafTemp++;
}
// pw.print(scaf.name);
if(tsw!=null && (sum>0 || !NONZERO_ONLY) && scaf.length>=minscaf){
if(TWOCOLUMN){
tsw.print(Tools.format("%s\t%.4f\n", scaf.name, sum/(double)scaf.length));
}else if(COUNT_GC){
long[] bc=scaf.basecount;
double gc=(bc[1]+bc[2])*1d/Data.max(1, bc[0]+bc[1]+bc[2]+bc[3]);
if(USE_COVERAGE_ARRAYS){
if(USE_WINDOW){
//Variable portion:
//"\tRead_GC\tMedian_fold\tStd_Dev\tUnder_X\n"
//\t%.4f\t%d\t%.2f\t%d\n
tsw.print(Tools.format("%s\t%.4f\t%d\t%.4f\t%.4f\t%d\t%d\t%d\t%.4f\t%d\t%.2f\t%d\n", scaf.name, sum/(double)scaf.length, scaf.length,
scaf.gc, covered*100d/scaf.length, covered, (scaf.readhits-scaf.readhitsMinus), scaf.readhitsMinus,
gc, median, stdev, underWindowAverage));
}else{
tsw.print(Tools.format("%s\t%.4f\t%d\t%.4f\t%.4f\t%d\t%d\t%d\t%.4f\t%d\t%.2f\n", scaf.name, sum/(double)scaf.length, scaf.length,
scaf.gc, covered*100d/scaf.length, covered, (scaf.readhits-scaf.readhitsMinus), scaf.readhitsMinus,
gc, median, stdev));
}
}else{
tsw.print(Tools.format("%s\t%.4f\t%d\t%.4f\t%.4f\t%d\t%d\t%d\t%.4f\n", scaf.name, sum/(double)scaf.length, scaf.length,
scaf.gc, covered*100d/scaf.length, covered, (scaf.readhits-scaf.readhitsMinus), scaf.readhitsMinus,
gc));
}
}else{
if(USE_COVERAGE_ARRAYS){
if(USE_WINDOW){
tsw.print(Tools.format("%s\t%.4f\t%d\t%.4f\t%.4f\t%d\t%d\t%d\t%d\t%.2f\t%d\n", scaf.name, sum/(double)scaf.length, scaf.length,
scaf.gc, covered*100d/scaf.length, covered, (scaf.readhits-scaf.readhitsMinus), scaf.readhitsMinus,
median, stdev, underWindowAverage));
}else{
tsw.print(Tools.format("%s\t%.4f\t%d\t%.4f\t%.4f\t%d\t%d\t%d\t%d\t%.2f\n", scaf.name, sum/(double)scaf.length, scaf.length,
scaf.gc, covered*100d/scaf.length, covered, (scaf.readhits-scaf.readhitsMinus), scaf.readhitsMinus,
median, stdev));
}
}else{
tsw.print(Tools.format("%s\t%.4f\t%d\t%.4f\t%.4f\t%d\t%d\t%d\n", scaf.name, sum/(double)scaf.length, scaf.length,
scaf.gc, covered*100d/scaf.length, covered, (scaf.readhits-scaf.readhitsMinus), scaf.readhitsMinus));
}
}
}
coveredBaseTemp+=covered;
}
if(strand==0){
scaffoldsWithCoverage1+=coveredScafTemp;
totalCoveredBases1+=coveredBaseTemp;
}else{
scaffoldsWithCoverage2+=coveredScafTemp;
totalCoveredBases2+=coveredBaseTemp;
}
if(tsw!=null){tsw.poisonAndWait();}
return hist==null ? null : hist.array;
}
/**
* Write a histogram of number of bases covered to each depth
* @param fname Output filename
* @param counts counts[X] stores the number of bases with coverage X
*/
public static void writeHist(String fname, long[] counts){
if(fname==null){return;}
assert(counts!=null) : "Can't write a histogram with null counts.";
ByteStreamWriter tsw=new ByteStreamWriter(fname, overwrite, false, false);
tsw.start();
if(printHeader){
if(headerPound){tsw.print('#');}
tsw.println("Coverage\tnumBases");
}
int max=0;
for(max=counts.length-1; max>0 && counts[max]==0; max--){}
for(int i=0; i<=max; i++){
long x=counts[i];
tsw.print(i);
tsw.print('\t');
tsw.println(x);
}
tsw.poisonAndWait();
}
/**
* Prints coverage in this format:
* scafname TAB position TAB coverage
* scafname TAB position TAB coverage
* @param fname Output filename
* @param list List of reference scaffolds
* @param deltaOnly Only write lines when coverage changes
* @param strand Only use coverage from reads mapped to this strand (0=plus, 1=minus)
*/
public void writeCoveragePerBase(String fname, ArrayList<Scaffold> list, boolean deltaOnly, int strand, int minscaf){
if(fname==null || (!STRANDED && strand>0)){return;}
if(verbose){outstream.println("Starting tsw "+fname);}
ByteStreamWriter tsw=new ByteStreamWriter(fname, overwrite, false, true);
if(verbose){outstream.println("Created tsw "+fname);}
tsw.start();
// if(verbose){outstream.println("Started tsw "+fname);}
if(printHeader){
if(headerPound){tsw.print('#');}
tsw.println("RefName\tPos\tCoverage");
}
for(Scaffold scaf : list){
int last=-1;
CoverageArray ca=(CoverageArray)(STRANDED && strand==1 ? scaf.obj1 : scaf.obj0);
if(scaf.length>=minscaf){
for(int i=0, len=scaf.length; i<len; i++){
int x=(ca==null ? 0 : ca.get(i));
if(!deltaOnly || x!=last){
if(x>0 || !NONZERO_ONLY){
tsw.print(scaf.name);
tsw.print('\t');
tsw.print(i);
tsw.print('\t');
tsw.println(x);
}
last=x;
}
}
}
}
if(verbose){outstream.println("Closing tsw "+fname);}
tsw.poisonAndWait();
if(verbose){outstream.println("Closed tsw "+fname);}
}
/**
* Prints coverage in this format, skipping zero-coverage positions:
* #scafname
* position TAB coverage
* position TAB coverage
* @param fname Output filename
* @param list List of reference scaffolds
* @param strand Only use coverage from reads mapped to this strand (0=plus, 1=minus)
*/
public void writeCoveragePerBaseConcise(String fname, ArrayList<Scaffold> list, int strand, int minscaf){
if(fname==null || (!STRANDED && strand>0)){return;}
if(verbose){outstream.println("Starting tsw "+fname);}
ByteStreamWriter tsw=new ByteStreamWriter(fname, overwrite, false, true);
tsw.start();
if(verbose){outstream.println("Started tsw "+fname);}
// tsw.print(pound+"RefName\tPos\tCoverage\n");
for(Scaffold scaf : list){
tsw.print('#');
tsw.println(scaf.name);
CoverageArray ca=(CoverageArray)(STRANDED && strand==1 ? scaf.obj1 : scaf.obj0);
if(scaf.length>=minscaf){
for(int i=0; i<scaf.length; i++){
int x=(ca==null ? 0 : ca.get(i));
if(x>0){
tsw.print(i);
tsw.print('\t');
tsw.println(x);
}
}
}
}
if(verbose){outstream.println("Closing tsw "+fname);}
tsw.poisonAndWait();
if(verbose){outstream.println("Closed tsw "+fname);}
// assert(false);
}
/**
* Prints coverage in this format, skipping zero-coverage positions:
* #scafname
* position TAB coverage
* position TAB coverage
* @param fname Output filename
* @param list List of reference scaffolds
* @param strand Only use coverage from reads mapped to this strand (0=plus, 1=minus)
*/
public void writeCoverageRanges(String fname, ArrayList<Scaffold> list, int strand, int minscaf){
if(fname==null || (!STRANDED && strand>0)){return;}
if(verbose){outstream.println("Starting tsw "+fname);}
ByteStreamWriter tsw=new ByteStreamWriter(fname, overwrite, false, true);
tsw.start();
if(verbose){outstream.println("Started tsw "+fname);}
// tsw.print(pound+"RefName\tPos\tCoverage\n");
for(Scaffold scaf : list){
tsw.print('#');
tsw.println(scaf.name);
CoverageArray ca=(CoverageArray)(STRANDED && strand==1 ? scaf.obj1 : scaf.obj0);
if(scaf.length>=minscaf){
int start=-1;
int prev=-1;
long sum=0;
for(int i=0; i<scaf.length; i++){
int x=(ca==null ? 0 : ca.get(i));
if(x>=minDepthToBeCovered){
if(prev<minDepthToBeCovered) {//open range
start=i;
}
sum+=x;
}else if(prev>=minDepthToBeCovered){//close range
float len=i-start+1;
tsw.print(start).print('-').print(i-1).tab().println(sum/len, 2);
sum=0;
}
prev=x;
}
if(prev>=minDepthToBeCovered){//close final range
float len=scaf.length-start;
tsw.print(start).print('-').print(scaf.length-1).tab().println(sum/len, 2);
sum=0;
}
}
}
if(verbose){outstream.println("Closing tsw "+fname);}
tsw.poisonAndWait();
if(verbose){outstream.println("Closed tsw "+fname);}
// assert(false);
}
/**
* Note. As written, this will truncate all trailing bases of each scaffold's length modulo binsize.
* For example, with binsize 1000, the last 500 bases of a 1500 base scaffold will be ignored.
* @param fname Output filename
* @param list List of reference scaffolds
* @param binsize Width of coverage bins in bp
* @param strand Only use coverage from reads mapped to this strand (0=plus, 1=minus)
*/
public static void writeCoveragePerBaseBinned(String fname, ArrayList<Scaffold> list, int binsize, int strand, int minscaf){
if(fname==null || (!STRANDED && strand>0)){return;}
TextStreamWriter tsw=new TextStreamWriter(fname, overwrite, false, false);
tsw.start();
if(printHeader){
String pound=(headerPound ? "#" : "");
if(calcCovStdev){
double[] stdev=standardDeviation(list, strand, minscaf);
if(stdev!=null){
tsw.print(pound+"Mean\t"+Tools.format("%.3f", stdev[0])+"\n");
tsw.print(pound+"STDev\t"+Tools.format("%.3f", stdev[1])+"\n");
}
}
tsw.print(pound+"RefName\tCov\tPos\tRunningPos\n");
}
long running=0;
final float invbin=1f/binsize;
for(Scaffold scaf : list){
if(scaf.length>=binsize && scaf.length>=minscaf){
CoverageArray ca=(CoverageArray)(STRANDED && strand==1 ? scaf.obj1 : scaf.obj0);
int lastPos=-1, nextPos=binsize-1;
long sum=0;
for(int i=0; i<scaf.length; i++){
int x=(ca==null ? 0 : ca.get(i));
sum+=x;
if(i>=nextPos){
if(sum>0 || !NONZERO_ONLY){
tsw.print(Tools.format("%s\t%.2f\t%d\t%d\n", scaf.name, sum*invbin, (i+1), running));
}
lastPos=i;
running+=binsize;
nextPos+=binsize;
sum=0;
}
}
}
}
tsw.poisonAndWait();
}
/**
* This version will NOT truncate all trailing bases of each scaffold's length modulo binsize.
* @param fname Output filename
* @param list List of reference scaffolds
* @param binsize Width of coverage bins in bp
* @param strand Only use coverage from reads mapped to this strand (0=plus, 1=minus)
*/
public static void writeCoveragePerBaseBinned2(String fname, ArrayList<Scaffold> list, int binsize, int strand, int minscaf){
if(fname==null || (!STRANDED && strand>0)){return;}
TextStreamWriter tsw=new TextStreamWriter(fname, overwrite, false, false);
tsw.start();
if(printHeader){
String pound=(headerPound ? "#" : "");
if(calcCovStdev){
double[] stdev=standardDeviationBinned(list, binsize, strand, minscaf);
if(stdev!=null){
tsw.print(pound+"Mean\t"+Tools.format("%.3f", stdev[0])+"\n");
tsw.print(pound+"STDev\t"+Tools.format("%.3f", stdev[1])+"\n");
}
}
tsw.print(pound+"RefName\tCov\tPos\tRunningPos\n");
}
long running=0;
for(Scaffold scaf : list){
CoverageArray ca=(CoverageArray)(STRANDED && strand==1 ? scaf.obj1 : scaf.obj0);
int lastPos=-1, nextPos=binsize-1;
long sum=0;
final int lim=scaf.length-1;
for(int i=0; i<scaf.length; i++){
int x=(ca==null ? 0 : ca.get(i));
sum+=x;
if(i>=nextPos || i==lim){
int bin=(i-lastPos);
if(scaf.length>=minscaf){
if(sum>0 || !NONZERO_ONLY){
tsw.print(Tools.format("%s\t%.2f\t%d\t%d\n", scaf.name, sum/(float)bin, (i+1), running));
}
}
running+=bin;
nextPos+=binsize;
lastPos=i;
sum=0;
}
}
}
tsw.poisonAndWait();
}
//Unsafe because it will fail if there are over 2 billion bins
public static double[] standardDeviationBinnedUnsafe(String fname, ArrayList<Scaffold> scaffolds, int binsize, int strand, int minscaf){
LongList list=new LongList();
for(Scaffold scaf : scaffolds){
CoverageArray ca=(CoverageArray)(STRANDED && strand==1 ? scaf.obj1 : scaf.obj0);
int lastPos=-1, nextPos=binsize-1;
long sum=0;
final int lim=scaf.length-1;
for(int i=0; i<scaf.length; i++){
int x=(ca==null ? 0 : ca.get(i));
sum+=x;
if(i>=nextPos || i==lim){
int bin=(i-lastPos);
if(scaf.length>=minscaf){
list.add((int)(10*(sum/(double)bin)));
}
nextPos+=binsize;
lastPos=i;
sum=0;
}
}
}
list.sort();
double mean=0.1*list.mean();
double median=0.1*list.median();
double mode=0.1*list.mode();
double stdev=0.1*list.stdev();
return new double[] {mean, median, mode, stdev};
}
public static double[] standardDeviationBinned(ArrayList<Scaffold> scaffolds, int binsize, int strand, int minscaf){
double totalSum=0;
long bins=0;
for(Scaffold scaf : scaffolds){
if(scaf.length>=minscaf){
CoverageArray ca=(CoverageArray)(STRANDED && strand==1 ? scaf.obj1 : scaf.obj0);
int lastPos=-1, nextPos=binsize-1;
long tempSum=0;
final int lim=scaf.length-1;
for(int i=0; i<scaf.length; i++){
int x=(ca==null ? 0 : ca.get(i));
tempSum+=x;
if(i>=nextPos || i==lim){
int bin=(i-lastPos);
double depth=(tempSum/(double)bin);
totalSum+=depth;
bins++;
nextPos+=binsize;
lastPos=i;
tempSum=0;
}
}
}
}
if(bins<1){return new double[] {0, 0};}
final double mean=totalSum/(double)bins;
double sumdev2=0;
for(Scaffold scaf : scaffolds){
if(scaf.length>=minscaf){
CoverageArray ca=(CoverageArray)(STRANDED && strand==1 ? scaf.obj1 : scaf.obj0);
int lastPos=-1, nextPos=binsize-1;
long tempSum=0;
final int lim=scaf.length-1;
for(int i=0; i<scaf.length; i++){
int x=(ca==null ? 0 : ca.get(i));
tempSum+=x;
if(i>=nextPos || i==lim){
int bin=(i-lastPos);
double depth=(tempSum/(double)bin);
double dev=mean-depth;
sumdev2+=(dev*dev);
nextPos+=binsize;
lastPos=i;
tempSum=0;
}
}
}
}
final double stdev=Math.sqrt(sumdev2/bins);
return new double[] {mean, stdev};
}
public static double[] standardDeviation(ArrayList<Scaffold> scaffolds, int strand, int minscaf){
long totalSum=0, bins=0, scafsCounted=0;
for(Scaffold scaf : scaffolds){
// System.err.println(scaf.name+", "+scaf.length+"/"+minscaf);
if(scaf.length>=minscaf){
final CoverageArray ca=(CoverageArray)(STRANDED && strand==1 ? scaf.obj1 : scaf.obj0);
bins+=scaf.length;
scafsCounted++;
if(ca!=null) {
for(int i=0; i<scaf.length; i++){
int depth=(ca==null ? 0 : ca.get(i));
totalSum+=depth;
}
}
}
}
if(bins<1){return new double[] {0, 0};}
final double mean=totalSum/(double)bins;
double sumdev2=0;
// System.err.println("sum="+totalSum+", bins="+bins+", mean="+mean+", scaffolds="+scafsCounted);
for(Scaffold scaf : scaffolds){
if(scaf.length>=minscaf){
final CoverageArray ca=(CoverageArray)(STRANDED && strand==1 ? scaf.obj1 : scaf.obj0);
double sumTemp=0;
if(ca!=null) {
for(int i=0; i<scaf.length; i++){
int depth=(ca==null ? 0 : ca.get(i));
double dev=mean-depth;
sumTemp+=(dev*dev);
}
}else {
sumTemp+=(mean*mean*scaf.length);
}
sumdev2+=sumTemp;
}
}
final double stdev=Math.sqrt(sumdev2/bins);
// System.err.println("sumdev2="+sumdev2+", stdev="+stdev);
return new double[] {mean, stdev};
}
/**
* @param fname Output filename
* @param list List of reference scaffolds
* @param binsize Width of coverage bins in bp
* @param strand Only use coverage from reads mapped to this strand (0=plus, 1=minus)
*/
public static void writeCoveragePerBaseNormalized(String fname, ArrayList<Scaffold> list, double binsize, int strand, int minscaf){
if(fname==null || (!STRANDED && strand>0)){return;}
TextStreamWriter tsw=new TextStreamWriter(fname, overwrite, false, false);
tsw.start();
if(printHeader){
String pound=(headerPound ? "#" : "");
tsw.print(pound+"RefName\tBin\tCov\tPos\tRunningPos\n");
}
double running=0;
double invbin=1.0/binsize;
final double invbincount=1.0/NORMALIZE_LENGTH_BINS;
for(Scaffold scaf : list){
if(NORMALIZE_LENGTH_BINS>0){
binsize=scaf.length*invbincount;
invbin=1.0/binsize;
}
if(scaf.length>=binsize && scaf.length>=minscaf){
long max=-1;
final CoverageArray ca=(CoverageArray)(STRANDED && strand==1 ? scaf.obj1 : scaf.obj0);
double lastPos=-1, nextPos=binsize-1;
long sum=0;
if(NORMALIZE_COVERAGE){
for(int i=0; i<scaf.length; i++){
int x=(ca==null ? 0 : ca.get(i));
sum+=x;
if(i>=nextPos){
max=Tools.max(sum, max);
running+=binsize;
nextPos+=binsize;
sum=0;
}
}
lastPos=-1;
nextPos=binsize-1;
sum=0;
assert(max>-1) : max;
}
max=Tools.max(max, 1);
final double binmult=(NORMALIZE_COVERAGE ? 1d/max : invbin);
// assert(false) : NORMALIZE_COVERAGE+", "+binmult+", "+invbin+", "+max+", "+binsize;
final String formatString=NORMALIZE_COVERAGE ? "%s\t%d\t%.5f\t%d\t%d\n" : "%s\t%d\t%.2f\t%d\t%d\n";
int bin=1;
for(int i=0; i<scaf.length; i++){
int x=(ca==null ? 0 : ca.get(i));
sum+=x;
if(i>=nextPos){
// outstream.println(x+", "+i+", "+nextPos+", "+sum+", "+(sum*binmult));
tsw.print(String.format(formatString, scaf.name, bin, sum*binmult, (i+1), (long)running));
bin++;
lastPos=i;
running+=binsize;
nextPos+=binsize;
sum=0;
}
}
}
}
tsw.poisonAndWait();
}
/**
* @param fname Output filename
* @param list List of reference scaffolds
* @param binsize Width of coverage bins in bp
* @param strand Only use coverage from reads mapped to this strand (0=plus, 1=minus)
*/
public static void writeCoveragePerBaseNormalizedOverall(String fname, ArrayList<Scaffold> list, double binsize, int strand, int minscaf){
if(fname==null || (!STRANDED && strand>0)){return;}
assert(NORMALIZE_LENGTH_BINS>0) : "Must set 'normalizebins' flag to a positive integer.";
double running=0;
double invbin=1.0/binsize;
long usedScafs=0;
final double invbincount=1.0/NORMALIZE_LENGTH_BINS;
double[] normalized=new double[NORMALIZE_LENGTH_BINS+1];
double[] absolute=new double[NORMALIZE_LENGTH_BINS+1];
for(Scaffold scaf : list){
if(NORMALIZE_LENGTH_BINS>0){
binsize=scaf.length*invbincount;
invbin=1.0/binsize;
}
if(scaf.length>=binsize && scaf.length>=minscaf){
usedScafs++;
if(scaf.readhits>0){
long max=-1;
final CoverageArray ca=(CoverageArray)(STRANDED && strand==1 ? scaf.obj1 : scaf.obj0);
double lastPos=-1, nextPos=binsize-1;
long sum=0;
{
for(int i=0; i<scaf.length; i++){
int x=(ca==null ? 0 : ca.get(i));
sum+=x;
if(i>=nextPos){
max=Tools.max(sum, max);
running+=binsize;
nextPos+=binsize;
sum=0;
}
}
lastPos=-1;
nextPos=binsize-1;
sum=0;
assert(max>-1) : max;
}
max=Tools.max(max, 1);
final double binmult=1d/max;
// assert(false) : NORMALIZE_COVERAGE+", "+binmult+", "+invbin+", "+max+", "+binsize;
int bin=1;
for(int i=0; i<scaf.length; i++){
int x=(ca==null ? 0 : ca.get(i));
sum+=x;
if(i>=nextPos){
normalized[bin]+=(sum*binmult);
absolute[bin]+=(sum*invbin);
bin++;
lastPos=i;
running+=binsize;
nextPos+=binsize;
sum=0;
}
}
}
}
}
TextStreamWriter tsw=new TextStreamWriter(fname, overwrite, false, false);
tsw.start();
if(printHeader){
String pound=(headerPound ? "#" : "");
tsw.print(pound+"RefName\tBin\tAbsCov\tNormCov\n");
}
double invScafs=1d/Tools.max(1, usedScafs);
final double maxNorm=Tools.max(normalized);
final double normMult=1/maxNorm;
for(int bin=1; bin<normalized.length; bin++){
// assert((absolute[bin]*invScafs)!=Double.NaN && (normalized[bin]*invScafs)!=Double.NaN) : invScafs+", "+absolute[bin]+", "+normalized[bin];
// assert(false) : invScafs+", "+absolute[bin]+", "+normalized[bin]+", "+(absolute[bin]*invScafs)+", "+(normalized[bin]*invScafs);
tsw.print(Tools.format("%s\t%d\t%.5f\t%.5f\n", "all", bin, absolute[bin]*invScafs, normalized[bin]*normMult));
}
tsw.poisonAndWait();
}
/**
* Write RPKM statistics.
*/
public static void writeRPKM(String out, String in1, String in2, long readsIn, boolean printNonZeroOnly, ArrayList<Scaffold> list){
if(out==null){return;}
final TextStreamWriter tsw=new TextStreamWriter(out, overwrite, false, false);
tsw.start();
/* Count mapped reads */
long mappedReads=0;
long mappedFrags=0;
for(Scaffold scaf : list){
mappedReads+=scaf.readhits;
mappedFrags+=scaf.fraghits;
}
mappedFrags/=2;
/* Print header */
tsw.print("#File\t"+(in1==null ? "" : in1)+(in2==null ? "" : "\t"+in2)+"\n");
tsw.print(Tools.format("#Reads\t%d\n",readsIn));
tsw.print(Tools.format("#Mapped\t%d\n",mappedReads));
tsw.print(Tools.format("#RefSequences\t%d\n",list.size()));
tsw.print("#Name\tLength\tBases\tCoverage\tReads\tRPKM\tFrags\tFPKM\n");
final float readMult=1000000000f/Tools.max(1, mappedReads);
final float fragMult=1000000000f/Tools.max(1, mappedFrags);
/* Print data */
for(final Scaffold scaf : list){
final long reads=scaf.readhits;
final long frags=scaf.fraghits/2;
final long bases=scaf.basehits;
final String s=scaf.name;
final int len=scaf.length;
final double invlen=1.0/Tools.max(1, len);
final double readMult2=readMult*invlen;
final double fragMult2=fragMult*invlen;
if(reads>0 || !printNonZeroOnly){
tsw.print(Tools.format("%s\t%d\t%d\t%.4f\t%d\t%.4f\t%d\t%.4f\n",s,len,bases,bases*invlen,reads,reads*readMult2,frags,frags*fragMult2));
}
}
tsw.poisonAndWait();
}
/*--------------------------------------------------------------*/
/*---------------- Fields ----------------*/
/*--------------------------------------------------------------*/
/** The list of all scaffolds */
private ArrayList<Scaffold> list;
/** Maps names to scaffolds */
private HashMap<String, Scaffold> table;
/** Converts BBMap index coordinates to scaffold coordinates */
private final ScaffoldCoordinates coords=new ScaffoldCoordinates(), coords2=new ScaffoldCoordinates();
/** Mapping program name */
private String program=null;
/** Mapping program version */
private String version=null;
//Inputs
/** Primary input file (typically sam) */
public String in1=null;
/** Secondary input file (typically for coverage directly from BBMap) */
public String in2=null;
public ArrayList<String> covIn=new ArrayList<String>();
/** Optional, for calculating GC */
public String reference=null;
public String orffasta=null;
//Outputs
/** Stream unaltered sam input to this output */
public String outsam=null;
/** Coverage statistics, one line per scaffold */
public String covstats=null;
public String outorf=null;
/** Coverage histogram, one line per depth and one point per base */
public String histogram=null;
/** Coverage with one line per base */
public String basecov=null;
/** Coverage with one file per scaffold */
public String basecov_ps=null;
/** Coverage with one line per bin */
public String bincov=null;
/** Coverage with one line per bin, normalized by length and/or height */
public String normcov=null;
/** Coverage with one line per bin, normalized by length and/or height, for combined reference */
public String normcovOverall=null;
/** rpkm/fpkm output, similar to Seal */
public String outrpkm=null;
public String rangecov=null;
/** Typically indicates that a header line was encountered in an unexpected place, e.g. with concatenated sam files. */
private boolean error=false;
private boolean warned=false;
private final boolean EA=Shared.EA();
/** Total length of reference */
public long refBases=0;
public long mappedBases=0;
public long mappedNonClippedBases=0;
public long mappedBasesWithDels=0;
public long mappedReads=0;
public long properPairs=0;
public long readsProcessed=0;
public long basesProcessed=0;
public long kmersProcessed=0;
public long mappedKmers=0;
public double correctKmers=0;
public long totalCoveredBases1=0;
public long totalCoveredBases2=0;
public long scaffoldsWithCoverage1=0;
public long scaffoldsWithCoverage2=0;
public long totalScaffolds=0;
public int k=0;
//Don't reset these variables when clearing.
public long maxReads=-1;
public int initialScaffolds=4096;
public int binsize=1000;
public boolean bits32=false;
public int minMapq=0;
public boolean useStreamer=true;
public int streamerThreads=2;
public int minDepthToBeCovered=1;
private boolean qtrimLeft=false;
private boolean qtrimRight=false;
private float trimq=-1;
private final float trimE;
private int border=0;
/** Don't print coverage info for scaffolds shorter than this */
public int minscaf=0;
public HashMap<String, SamLine> pairTable=new HashMap<String, SamLine>();
public PrintStream outstream=System.err;
private boolean errorState=false;
private final LineParser1 lp=new LineParser1('\t');
/*--------------------------------------------------------------*/
/*---------------- Static Fields ----------------*/
/*--------------------------------------------------------------*/
/** Permission to overwrite existing files */
public static boolean overwrite=true;
/** Permission to append to existing files */
public static boolean append=false;
/** Print verbose log messages */
public static boolean verbose=false;
/** Print the arguments to main */
public static boolean printCommand=true;
/** Print headers in output files */
public static boolean printHeader=true;
/** Prepend '#' symbol to header lines */
public static boolean headerPound=true;
/** Calculate standard deviation of coverage */
public static boolean calcCovStdev=true;
/** Window size to use when calculating average coverage,
* for detecting contiguous low-coverage areas */
public static int LOW_COV_WINDOW=500;
/** Min average coverage to not be classified as low-depth */
public static double LOW_COV_DEPTH=5;
/** Print number of bases below a certain average coverage in a window */
public static boolean USE_WINDOW=false;
/** Track base composition of reads covering each scaffold */
public static boolean COUNT_GC=true;
/** Output in 2-column format ("#ID\tAvg_fold\n") */
public static boolean TWOCOLUMN=false;
/** Track coverage for strands independently */
public static boolean STRANDED=false;
/** Add scaffold information from the reference (in addition to sam header) */
public static boolean ADD_FROM_REF=true;
/** Add scaffold information from reads mapped to unknown scaffolds (in addition to sam header) */
public static boolean ADD_FROM_READS=false;
/** Only print scaffolds with nonzero coverage */
public static boolean NONZERO_ONLY=false;
/** Store coverage info in numeric arrays */
public static boolean USE_COVERAGE_ARRAYS=true;
/** Store coverage info in bitsets */
public static boolean USE_BITSETS=false;
/** Only print lines when coverage changes (for compatibility with Jigsaw) */
public static boolean DELTA_ONLY=false;
/** Process secondary alignments */
public static boolean USE_SECONDARY=true;
/** Include coverage of unsequenced middle portion of pairs */
public static boolean PHYSICAL_COVERAGE=false;
/** Use 'tlen' field when calculating physical coverage */
public static boolean USE_TLEN=true;
/** Abort on error; otherwise, errors may be ignored */
public static boolean ABORT_ON_ERROR=true;
/** Print coverage for the last bin of a scaffold, even if it is shorter than binsize */
public static boolean KEEP_SHORT_BINS=true;
/** Only track coverage for start location */
public static boolean START_ONLY=false;
/** Only track coverage for stop location */
public static boolean STOP_ONLY=false;
/** This appears to be the same as nonzeroonly... */
public static boolean CONCISE=false;
/** Normalize coverage by expression contig coverage as a fraction of its max coverage */
public static boolean NORMALIZE_COVERAGE=false;
/** Normalize contig length by binning into this many bins per contig */
public static int NORMALIZE_LENGTH_BINS=100;
/** Include soft-clipped bases in coverage */
public static boolean INCLUDE_SOFT_CLIP=false;
/** Include deletions/introns in coverage */
public static boolean INCLUDE_DELETIONS=true;
/** Include reads flagged as duplicates in coverage */
public static boolean INCLUDE_DUPLICATES=true;
public static boolean KEY_VALUE;
/** Translation array for tracking base counts */
private static final byte[] charToNum=AssemblyStats2.makeCharToNum();
private static final int NOTHING_MODE=0, BITSET_MODE=1, ARRAY_MODE=2;
}
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