package edu.mit.broad.picard.sam; import edu.mit.broad.picard.metrics.MetricBase; import edu.mit.broad.picard.util.Histogram; /** * Metrics that are calculated during the process of marking duplicates * within a stream of SAMRecords. */ public class DuplicationMetrics extends MetricBase { /** The number of mapped reads examined which did not have a mapped mate pair. */ public long UNPAIRED_READS_EXAMINED; /** The number of mapped read pairs examined. */ public long READ_PAIRS_EXAMINED; /** The total number of unmapped reads examined. */ public long UNMAPPED_READS; /** The number of fragments that were marked as duplicates. */ public long UNPAIRED_READ_DUPLICATES; /** The number of read pairs that were marked as duplicates. */ public long READ_PAIR_DUPLICATES; /** The percentage of mapped sequence that is marked as duplicate. */ public Double PERCENT_DUPLICATION; /** The estimated number of unique molecules in the library based on PE duplication. */ public Long ESTIMATED_LIBRARY_SIZE; /** * Fills in the ESTIMATED_LIBRARY_SIZE based on the paired read data examined where * possible and the PERCENT_DUPLICATION. */ public void calculateDerivedMetrics() { if (READ_PAIRS_EXAMINED > 0) { // Following code "borrowed" from CRD codebase long n = READ_PAIRS_EXAMINED; long c = READ_PAIRS_EXAMINED - READ_PAIR_DUPLICATES; double m = 1.0, M = 100.0; if (c >= n || f(m*c, c, n) <= 0) { throw new IllegalStateException("Invalid values for pairs and unique pairs: " + n + ", " + c); } while( f(M*c, c, n) >= 0 ) M *= 10.0; for (int i=0; i<40; i++ ) { double r = (m+M)/2.0; double u = f( r * c, c, n ); if ( u == 0 ) break; else if ( u > 0 ) m = r; else if ( u < 0 ) M = r; } this.ESTIMATED_LIBRARY_SIZE = (long) (c * (m+M)/2.0); } PERCENT_DUPLICATION = (UNPAIRED_READ_DUPLICATES + READ_PAIR_DUPLICATES *2) /(double) (UNPAIRED_READS_EXAMINED + READ_PAIRS_EXAMINED *2); } /** Method that is used in the computation of estimated library size. */ private double f(double x, double c, double n) { return c/x - 1 + Math.exp(-n/x); } /** * Estimates the ROI (return on investment) that one would see if a library was sequenced to * x higher coverage than the observed coverage. * * @param estimatedLibrarySize the estimated number of molecules in the library * @param x the multiple of sequencing to be simulated (i.e. how many X sequencing) * @param pairs the number of pairs observed in the actual sequencing * @param uniquePairs the number of unique pairs observed in the actual sequencing * @return a number z <= x that estimates if you had pairs*x as your sequencing then you * would observe uniquePairs*z unique pairs. */ private double estimateRoi(long estimatedLibrarySize, double x, long pairs, long uniquePairs) { return estimatedLibrarySize * ( 1 - Math.exp(-(x*pairs)/estimatedLibrarySize) ) / uniquePairs; } /** * Calculates a histogram using the estimateRoi method to estimate the effective yield * doing x sequencing for x=1..10. */ public Histogram calculateRoiHistogram() { if (ESTIMATED_LIBRARY_SIZE == null) { try { calculateDerivedMetrics(); } catch (IllegalStateException ise) { return null; } } long uniquePairs = READ_PAIRS_EXAMINED - READ_PAIR_DUPLICATES; Histogram histo = new Histogram(); for (double x=1; x<=10; x+=1) { histo.increment(x, estimateRoi(ESTIMATED_LIBRARY_SIZE, x, READ_PAIRS_EXAMINED, uniquePairs)); } return histo; } // Main method used for debugging the derived metrics // public static void main(String[] args) { // DuplicationMetrics m = new DuplicationMetrics(); // m.PAIRS_EXAMINED = Integer.parseInt(args[0]); // m.DUPLICATE_PAIRS = m.PAIRS_EXAMINED - Integer.parseInt(args[1]); // m.calculateDerivedMetrics(); // System.out.println("Percent Duplication: " + m.PERCENT_DUPLICATION); // System.out.println("Est. Library Size : " + m.ESTIMATED_LIBRARY_SIZE); // System.out.println(m.calculateRoiHistogram()); // } }