#!/usr/bin/env k8 var paftools_version = 'r713'; /***************************** ***** Library functions ***** *****************************/ /******************************* * Command line option parsing * *******************************/ var getopt = function(args, ostr) { var oli; // option letter list index if (typeof(getopt.place) == 'undefined') getopt.ind = 0, getopt.arg = null, getopt.place = -1; if (getopt.place == -1) { // update scanning pointer if (getopt.ind >= args.length || args[getopt.ind].charAt(getopt.place = 0) != '-') { getopt.place = -1; return null; } if (getopt.place + 1 < args[getopt.ind].length && args[getopt.ind].charAt(++getopt.place) == '-') { // found "--" ++getopt.ind; getopt.place = -1; return null; } } var optopt = args[getopt.ind].charAt(getopt.place++); // character checked for validity if (optopt == ':' || (oli = ostr.indexOf(optopt)) < 0) { if (optopt == '-') return null; // if the user didn't specify '-' as an option, assume it means null. if (getopt.place < 0) ++getopt.ind; return '?'; } if (oli+1 >= ostr.length || ostr.charAt(++oli) != ':') { // don't need argument getopt.arg = null; if (getopt.place < 0 || getopt.place >= args[getopt.ind].length) ++getopt.ind, getopt.place = -1; } else { // need an argument if (getopt.place >= 0 && getopt.place < args[getopt.ind].length) getopt.arg = args[getopt.ind].substr(getopt.place); else if (args.length <= ++getopt.ind) { // no arg getopt.place = -1; if (ostr.length > 0 && ostr.charAt(0) == ':') return ':'; return '?'; } else getopt.arg = args[getopt.ind]; // white space getopt.place = -1; ++getopt.ind; } return optopt; } /*********************** * Interval operations * ***********************/ Interval = {}; Interval.sort = function(a) { if (typeof a[0] == 'number') a.sort(function(x, y) { return x - y }); else a.sort(function(x, y) { return x[0] != y[0]? x[0] - y[0] : x[1] - y[1] }); } Interval.merge = function(a, sorted) { if (typeof sorted == 'undefined') sorted = true; if (!sorted) Interval.sort(a); var k = 0; for (var i = 1; i < a.length; ++i) { if (a[k][1] >= a[i][0]) a[k][1] = a[k][1] > a[i][1]? a[k][1] : a[i][1]; else a[++k] = a[i].slice(0); } a.length = k + 1; } Interval.index_end = function(a, sorted) { if (a.length == 0) return; if (typeof sorted == 'undefined') sorted = true; if (!sorted) Interval.sort(a); a[0].push(0); var k = 0, k_en = a[0][1]; for (var i = 1; i < a.length; ++i) { if (k_en <= a[i][0]) { for (++k; k < i; ++k) if (a[k][1] > a[i][0]) break; k_en = a[k][1]; } a[i].push(k); } } Interval.find_intv = function(a, x) { var left = -1, right = a.length; if (typeof a[0] == 'number') { while (right - left > 1) { var mid = left + ((right - left) >> 1); if (a[mid] > x) right = mid; else if (a[mid] < x) left = mid; else return mid; } } else { while (right - left > 1) { var mid = left + ((right - left) >> 1); if (a[mid][0] > x) right = mid; else if (a[mid][0] < x) left = mid; else return mid; } } return left; } Interval.find_ovlp = function(a, st, en) { if (a.length == 0 || st >= en) return []; var l = Interval.find_intv(a, st); var k = l < 0? 0 : a[l][a[l].length - 1]; var b = []; for (var i = k; i < a.length; ++i) { if (a[i][0] >= en) break; else if (st < a[i][1]) b.push(a[i]); } return b; } /********************************** * Reverse and reverse complement * **********************************/ Bytes.prototype.reverse = function() { for (var i = 0; i < this.length>>1; ++i) { var tmp = this[i]; this[i] = this[this.length - i - 1]; this[this.length - i - 1] = tmp; } } // reverse complement a DNA string Bytes.prototype.revcomp = function() { if (Bytes.rctab == null) { var s1 = 'WSATUGCYRKMBDHVNwsatugcyrkmbdhvn'; var s2 = 'WSTAACGRYMKVHDBNwstaacgrymkvhdbn'; Bytes.rctab = []; for (var i = 0; i < 256; ++i) Bytes.rctab[i] = 0; for (var i = 0; i < s1.length; ++i) Bytes.rctab[s1.charCodeAt(i)] = s2.charCodeAt(i); } for (var i = 0; i < this.length>>1; ++i) { var tmp = this[this.length - i - 1]; this[this.length - i - 1] = Bytes.rctab[this[i]]; this[i] = Bytes.rctab[tmp]; } if (this.length&1) this[this.length>>1] = Bytes.rctab[this[this.length>>1]]; } /******************** ***** paftools ***** ********************/ /***************** * Miscellaneous * *****************/ // liftover function paf_liftover(args) { function read_bed(fn, to_merge) { if (fn == null) return null; if (typeof to_merge == 'undefined') to_merge = true; var file = fn == '-'? new File() : new File(fn); var buf = new Bytes(); var bed = {}; while (file.readline(buf) >= 0) { var t = buf.toString().split("\t"); if (bed[t[0]] == null) bed[t[0]] = []; bed[t[0]].push([parseInt(t[1]), parseInt(t[2])]); } buf.destroy(); file.close(); for (var chr in bed) { Interval.sort(bed[chr]); if (to_merge) Interval.merge(bed[chr], true); Interval.index_end(bed[chr], true); } return bed; } var re_cigar = /(\d+)([MID])/g, re_tag = /^(\S\S):([AZif]):(\S+)$/; var c, to_merge = false, min_mapq = 5, min_len = 50000, max_div = 2.0; var re = /(\d+)([MID])/g; while ((c = getopt(args, "mq:l:d:")) != null) { if (c == 'm') to_merge = true; else if (c == 'q') min_mapq = parseInt(getopt.arg); else if (c == 'l') min_len = parseInt(getopt.arg); else if (c == 'd') max_div = parseFloat(getopt.arg); } if (args.length - getopt.ind < 2) { print("Usage: paftools.js liftover [options] "); print("Options:"); print(" -q INT min mapping quality [" + min_mapq + "]"); print(" -l INT min alignment length [" + min_len + "]"); print(" -d FLOAT max sequence divergence (>=1 to disable) [1]"); exit(1); } var bed = read_bed(args[getopt.ind+1], to_merge); var file = args[getopt.ind] == '-'? new File() : new File(args[getopt.ind]); var buf = new Bytes(); while (file.readline(buf) >= 0) { var t = buf.toString().split("\t"); if (bed[t[0]] == null) continue; // sequence not present in BED; skip // parse tp and cg tags var m, tp = null, cg = null; for (var i = 12; i < t.length; ++i) { if ((m = re_tag.exec(t[i])) != null) { if (m[1] == 'tp') tp = m[3]; else if (m[1] == 'cg') cg = m[3]; } } if (tp != 'P' && tp != 'I') continue; // only process primary alignments if (cg == null) throw Error("unable to find the 'cg' tag"); // filter out bad alignments and check overlaps for (var i = 1; i <= 3; ++i) t[i] = parseInt(t[i]); for (var i = 6; i <= 11; ++i) t[i] = parseInt(t[i]); if (t[11] < min_mapq || t[10] < min_len) continue; var regs = Interval.find_ovlp(bed[t[0]], t[2], t[3]); if (regs.length == 0) continue; // not overlapping any regions in input BED if (max_div >= 0.0 && max_div < 1.0) { var n_gaps = 0, n_opens = 0; while ((m = re_cigar.exec(cg)) != null) if (m[2] == 'I' || m[2] == 'D') n_gaps += parseInt(m[1]), ++n_opens; var n_mm = t[10] - t[9] - n_gaps; var n_diff2 = n_mm + n_opens; if (n_diff2 / (n_diff2 + t[9]) > max_div) continue; } // extract start and end positions var a = [], r = [], strand = t[4]; for (var i = 0; i < regs.length; ++i) { var s = regs[i][0], e = regs[i][1]; if (strand == '+') { a.push([s, 0, i, -2]); a.push([e - 1, 1, i, -2]); } else { a.push([t[1] - e, 0, i, -2]); a.push([t[1] - s - 1, 1, i, -2]); } r.push([-2, -2]); } a.sort(function(x, y) { return x[0] - y[0] }); // lift start/end positions var k = 0, x = t[7], y = strand == '+'? t[2] : t[1] - t[3]; while ((m = re_cigar.exec(cg)) != null) { // TODO: be more careful about edge cases var len = parseInt(m[1]); if (m[2] == 'D') { // do nothing for D x += len; continue; } while (k < a.length && a[k][0] < y) ++k; // skip out-of-range positions for (var i = k; i < a.length; ++i) { if (y <= a[i][0] && a[i][0] < y + len) a[i][3] = m[2] == 'M'? x + (a[i][0] - y) : x; else break; } y += len; if (m[2] == 'M') x += len; } if (x != t[8] || (strand == '+' && y != t[3]) || (strand == '-' && y != t[1] - t[2])) throw Error("CIGAR is inconsistent with mapping coordinates"); // generate result for (var i = 0; i < a.length; ++i) { if (a[i][1] == 0) r[a[i][2]][0] = a[i][3]; else r[a[i][2]][1] = a[i][3] + 1; // change to half-close-half-open } for (var i = 0; i < r.length; ++i) { var name = [t[0], regs[i][0], regs[i][1]].join("_"); if (r[i][0] < 0) name += "_t5", r[i][0] = t[7]; if (r[i][1] < 0) name += "_t3", r[i][1] = t[8]; print(t[5], r[i][0], r[i][1], name, 0, strand); } } buf.destroy(); file.close(); } // variant calling function paf_call(args) { var re_cs = /([:=*+-])(\d+|[A-Za-z]+)/g; var c, min_cov_len = 10000, min_var_len = 50000, gap_thres = 50, min_mapq = 5; while ((c = getopt(args, "l:L:g:q:B:")) != null) { if (c == 'l') min_cov_len = parseInt(getopt.arg); else if (c == 'L') min_var_len = parseInt(getopt.arg); else if (c == 'g') gap_thres = parseInt(getopt.arg); else if (c == 'q') min_mapq = parseInt(getopt.arg); } if (args.length == getopt.ind) { print("Usage: sort -k6,6 -k8,8n | paftools.js call [options] -"); print("Options:"); print(" -l INT min alignment length to compute coverage ["+min_cov_len+"]"); print(" -L INT min alignment length to call variants ["+min_var_len+"]"); print(" -q INT min mapping quality ["+min_mapq+"]"); print(" -g INT short/long gap threshold (for statistics only) ["+gap_thres+"]"); exit(1); } var file = args[getopt.ind] == '-'? new File() : new File(args[getopt.ind]); var buf = new Bytes(); var tot_len = 0, n_sub = [0, 0, 0], n_ins = [0, 0, 0, 0], n_del = [0, 0, 0, 0]; function count_var(o) { if (o[3] > 1) return; if (o[5] == '-' && o[6] == '-') return; if (o[5] == '-') { // insertion var l = o[6].length; if (l == 1) ++n_ins[0]; else if (l == 2) ++n_ins[1]; else if (l < gap_thres) ++n_ins[2]; else ++n_ins[3]; } else if (o[6] == '-') { // deletion var l = o[5].length; if (l == 1) ++n_del[0]; else if (l == 2) ++n_del[1]; else if (l < gap_thres) ++n_del[2]; else ++n_del[3]; } else { ++n_sub[0]; var s = o[5] + o[6]; if (s == 'ag' || s == 'ga' || s == 'ct' || s == 'tc') ++n_sub[1]; else ++n_sub[2]; } } var a = [], out = []; var c1_ctg = null, c1_start = 0, c1_end = 0, c1_counted = false, c1_len = 0; while (file.readline(buf) >= 0) { var line = buf.toString(); if (!/\ts2:i:/.test(line)) continue; // skip secondary alignments var m, t = line.split("\t", 12); for (var i = 6; i <= 11; ++i) t[i] = parseInt(t[i]); if (t[10] < min_cov_len || t[11] < min_mapq) continue; print(t[0], t[7], t[8], c1_start, c1_end); for (var i = 1; i <= 3; ++i) t[i] = parseInt(t[i]); var ctg = t[5], x = t[7], end = t[8]; var query = t[0], rev = (t[4] == '-'), y = rev? t[3] : t[2]; // compute regions covered by 1 contig if (ctg != c1_ctg || x >= c1_end) { if (c1_counted && c1_end > c1_start) { c1_len += c1_end - c1_start; print('R', c1_ctg, c1_start, c1_end); } c1_ctg = ctg, c1_start = x, c1_end = end; c1_counted = (t[10] >= min_var_len); } else if (end > c1_end) { // overlap if (c1_counted && x > c1_start) { c1_len += x - c1_start; print('R', c1_ctg, c1_start, x); } c1_start = c1_end, c1_end = end; c1_counted = (t[10] >= min_var_len); } else if (end > c1_start) { // contained if (c1_counted && x > c1_start) { c1_len += x - c1_start; print('R', c1_ctg, c1_start, x); } c1_start = end; } // else, the alignment precedes the cov1 region; do nothing // output variants ahead of this alignment while (out.length) { if (out[0][0] != ctg || out[0][2] <= x) { count_var(out[0]); print('V', out[0].join("\t")); out.shift(); } else break; } // update coverage for (var i = 0; i < out.length; ++i) if (out[i][1] >= x && out[i][2] <= end) ++out[i][3]; // drop alignments that don't overlap with the current one var k = 0; for (var i = 0; i < a.length; ++i) if (a[0][0] == ctg && a[0][2] > x) a[k++] = a[i]; a.length = k; // core loop if (t[10] >= min_var_len) { if ((m = /\tcs:Z:(\S+)/.exec(line)) == null) continue; // no cs tag var cs = m[1]; var blen = 0, n_diff = 0; tot_len += t[10]; while ((m = re_cs.exec(cs)) != null) { var cov = 1; if (m[1] == '*' || m[1] == '+' || m[1] == '-') for (var i = 0; i < a.length; ++i) if (a[0][2] > x) ++cov; var qs, qe; if (m[1] == '=' || m[1] == ':') { var l = m[1] == '='? m[2].length : parseInt(m[2]); if (rev) y -= l; else y += l; x += l, blen += l; } else if (m[1] == '*') { if (rev) qs = y - 1, qe = y, --y; else qs = y, qe = y + 1, ++y; var br = m[2].charAt(0), bq = m[2].charAt(1); if (br != 'n' && bq != 'n') { // don't call a SNP if there is an ambiguous base out.push([t[5], x, x+1, cov, t[11], br, bq, query, qs, qe, rev? '-' : '+']); ++n_diff; } ++x, ++blen; } else if (m[1] == '+') { var l = m[2].length; if (rev) qs = y - l, qe = y, y -= l; else qs = y, qe = y + l, y += l; out.push([t[5], x, x, cov, t[11], '-', m[2], query, qs, qe, rev? '-' : '+']); ++blen, ++n_diff; } else if (m[1] == '-') { var l = m[2].length; out.push([t[5], x, x + l, cov, t[11], m[2], '-', query, y, y, rev? '-' : '+']); x += l, ++blen, ++n_diff; } } } a.push([t[5], t[7], t[8]]); } if (c1_counted && c1_end > c1_start) { c1_len += c1_end - c1_start; print('R', c1_ctg, c1_start, c1_end); } while (out.length) { count_var(out[0]); print('V', out[0].join("\t")); out.shift(); } //warn(tot_len + " alignment columns considered in calling"); warn(c1_len + " reference bases covered by exactly one contig"); warn(n_sub[0] + " substitutions; ts/tv = " + (n_sub[1]/n_sub[2]).toFixed(3)); warn(n_del[0] + " 1bp deletions"); warn(n_ins[0] + " 1bp insertions"); warn(n_del[1] + " 2bp deletions"); warn(n_ins[1] + " 2bp insertions"); warn(n_del[2] + " [3,"+gap_thres+") deletions"); warn(n_ins[2] + " [3,"+gap_thres+") insertions"); warn(n_del[3] + " >="+gap_thres+" deletions"); warn(n_ins[3] + " >="+gap_thres+" insertions"); buf.destroy(); file.close(); } function paf_stat(args) { var c, gap_out_len = null; while ((c = getopt(args, "l:")) != null) if (c == 'l') gap_out_len = parseInt(getopt.arg); if (getopt.ind == args.length) { print("Usage: paftools.js stat [-l gapOutLen] |"); exit(1); } var buf = new Bytes(); var file = args[getopt.ind] == '-'? new File() : new File(args[getopt.ind]); var re = /(\d+)([MIDSHNX=])/g; var lineno = 0, n_pri = 0, n_2nd = 0, n_seq = 0, n_cigar_64k = 0, l_tot = 0, l_cov = 0; var n_gap = [[0, 0, 0, 0, 0, 0], [0, 0, 0, 0, 0, 0]]; function cov_len(regs) { regs.sort(function(a,b) {return a[0]-b[0]}); var st = regs[0][0], en = regs[0][1], l = 0; for (var i = 1; i < regs.length; ++i) { if (regs[i][0] < en) en = en > regs[i][1]? en : regs[i][1]; else l += en - st, st = regs[i][0], en = regs[i][1]; } l += en - st; return l; } var last = null, last_qlen = null, regs = []; while (file.readline(buf) >= 0) { var line = buf.toString(); ++lineno; if (line.charAt(0) != '@') { var t = line.split("\t", 12); var m, rs, cigar = null, is_pri = false, is_sam = false, is_rev = false, tname = null; var atlen = null, aqlen, qs, qe, mapq, ori_qlen; if (t[4] == '+' || t[4] == '-') { // PAF if (!/\ts2:i:\d+/.test(line)) { ++n_2nd; continue; } if ((m = /\tcg:Z:(\S+)/.exec(line)) != null) cigar = m[1]; if (cigar == null) { warn("WARNING: no CIGAR at line " + lineno); continue; } tname = t[5]; qs = parseInt(t[2]), qe = parseInt(t[3]); aqlen = qe - qs; is_rev = t[4] == '+'? false : true; rs = parseInt(t[7]); atlen = parseInt(t[8]) - rs; mapq = parseInt(t[11]); ori_qlen = parseInt(t[1]); } else { // SAM var flag = parseInt(t[1]); if ((flag & 4) || t[2] == '*' || t[5] == '*') continue; if (flag & 0x100) { ++n_2nd; continue; } cigar = t[5]; tname = t[2]; rs = parseInt(t[3]) - 1; mapq = parseInt(t[4]); aqlen = t[9].length; is_sam = true; is_rev = !!(flag&0x10); } ++n_pri; if (last != t[0]) { if (last != null) { l_tot += last_qlen; l_cov += cov_len(regs); } regs = []; ++n_seq, last = t[0]; } var M = 0, tl = 0, ql = 0, clip = [0, 0], n_cigar = 0, sclip = 0; while ((m = re.exec(cigar)) != null) { var l = parseInt(m[1]); ++n_cigar; if (m[2] == 'M' || m[2] == '=' || m[2] == 'X') { tl += l, ql += l, M += l; } else if (m[2] == 'I' || m[2] == 'D') { var type; if (l < 50) type = 0; else if (l < 100) type = 1; else if (l < 300) type = 2; else if (l < 400) type = 3; else if (l < 1000) type = 4; else type = 5; if (m[2] == 'I') ql += l, ++n_gap[0][type]; else tl += l, ++n_gap[1][type]; if (gap_out_len != null && l >= gap_out_len) print(t[0], ql, is_rev? '-' : '+', tname, rs + tl, m[2], l); } else if (m[2] == 'N') { tl += l; } else if (m[2] == 'S') { clip[M == 0? 0 : 1] = l, sclip += l; } else if (m[2] == 'H') { clip[M == 0? 0 : 1] = l; } } if (n_cigar > 65535) ++n_cigar_64k; if (ql + sclip != aqlen) warn("WARNING: aligned query length is inconsistent with CIGAR at line " + lineno + " (" + (ql+sclip) + " != " + aqlen + ")"); if (atlen != null && atlen != tl) warn("WARNING: aligned reference length is inconsistent with CIGAR at line " + lineno); if (is_sam) { qs = clip[is_rev? 1 : 0], qe = qs + ql; ori_qlen = clip[0] + ql + clip[1]; } regs.push([qs, qe]); last_qlen = ori_qlen; } } l_tot += last_qlen; l_cov += cov_len(regs); file.close(); buf.destroy(); if (gap_out_len == null) { print("Number of mapped sequences: " + n_seq); print("Number of primary alignments: " + n_pri); print("Number of secondary alignments: " + n_2nd); print("Number of primary alignments with >65535 CIGAR operations: " + n_cigar_64k); print("Number of bases in mapped sequences: " + l_tot); print("Number of mapped bases: " + l_cov); print("Number of insertions in [0,50): " + n_gap[0][0]); print("Number of insertions in [50,100): " + n_gap[0][1]); print("Number of insertions in [100,300): " + n_gap[0][2]); print("Number of insertions in [300,400): " + n_gap[0][3]); print("Number of insertions in [400,1000): " + n_gap[0][4]); print("Number of insertions in [1000,inf): " + n_gap[0][5]); print("Number of deletions in [0,50): " + n_gap[1][0]); print("Number of deletions in [50,100): " + n_gap[1][1]); print("Number of deletions in [100,300): " + n_gap[1][2]); print("Number of deletions in [300,400): " + n_gap[1][3]); print("Number of deletions in [400,1000): " + n_gap[1][4]); print("Number of deletions in [1000,inf): " + n_gap[1][5]); } } function paf_bedcov(args) { function read_bed(fn, to_merge, to_dedup) { var file = new File(fn); var buf = new Bytes(); var h = {}; while (file.readline(buf) >= 0) { var t = buf.toString().split("\t"); if (h[t[0]] == null) h[t[0]] = []; var bst = parseInt(t[1]); var ben = parseInt(t[2]); if (t.length >= 12 && /^\d+$/.test(t[9])) { t[9] = parseInt(t[9]); var sz = t[10].split(","); var st = t[11].split(","); for (var i = 0; i < t[9]; ++i) { st[i] = parseInt(st[i]); sz[i] = parseInt(sz[i]); h[t[0]].push([bst + st[i], bst + st[i] + sz[i], 0, 0, 0]); } } else { h[t[0]].push([bst, ben, 0, 0, 0]); } } buf.destroy(); file.close(); for (var chr in h) { if (to_merge) Interval.merge(h[chr], false); else if (to_dedup) Interval.dedup(h[chr], false); else Interval.sort(h[chr]); Interval.index_end(h[chr]); } return h; } var c, print_len = false, to_merge = true, to_dedup = false, fn_excl = null; while ((c = getopt(args, "pde:")) != null) { if (c == 'p') print_len = true; else if (c == 'd') to_dedup = true, to_merge = false; else if (c == 'e') fn_excl = getopt.arg; } if (args.length - getopt.ind < 2) { print("Usage: paftools.js bedcov [options] "); print("Options:"); print(" -e FILE exclude target regions (2nd file) overlapping BED FILE []"); print(" -p print number of covered bases for each target"); exit(1); } var excl = fn_excl != null? read_bed(fn_excl, true, false) : null; var target = read_bed(args[getopt.ind], to_merge, to_dedup); var file, buf = new Bytes(); var tot_len = 0, hit_len = 0; file = args[getopt.ind+1] != '-'? new File(args[getopt.ind+1]) : new File(); while (file.readline(buf) >= 0) { var t = buf.toString().split("\t"); var a = []; var bst = parseInt(t[1]); var ben = parseInt(t[2]); if (t.length >= 12 && /^\d+$/.test(t[9])) { // BED12 t[9] = parseInt(t[9]); var sz = t[10].split(","); var st = t[11].split(","); for (var i = 0; i < t[9]; ++i) { st[i] = parseInt(st[i]); sz[i] = parseInt(sz[i]); a.push([bst + st[i], bst + st[i] + sz[i], false]); } } else a.push([bst, ben, false]); // 3-column BED var feat_len = 0; for (var i = 0; i < a.length; ++i) { if (excl != null && excl[t[0]] != null) { var oe = Interval.find_ovlp(excl[t[0]], a[i][0], a[i][1]); if (oe.length > 0) continue; } a[i][2] = true; feat_len += a[i][1] - a[i][0]; } tot_len += feat_len; if (target[t[0]] == null) continue; var b = []; for (var i = 0; i < a.length; ++i) { if (!a[i][2]) continue; var o = Interval.find_ovlp(target[t[0]], a[i][0], a[i][1]); for (var j = 0; j < o.length; ++j) { var max_st = o[j][0] > a[i][0]? o[j][0] : a[i][0]; var min_en = o[j][1] < a[i][1]? o[j][1] : a[i][1]; b.push([max_st, min_en]); o[j][2] += min_en - max_st; ++o[j][3]; if (max_st == o[j][0] && min_en == o[j][1]) ++o[j][4]; } } // find the length covered var feat_hit_len = 0; if (b.length > 0) { b.sort(function(a,b) {return a[0]-b[0]}); var st = b[0][0], en = b[0][1]; for (var i = 1; i < b.length; ++i) { if (b[i][0] <= en) en = en > b[i][1]? en : b[i][1]; else feat_hit_len += en - st, st = b[i][0], en = b[i][1]; } feat_hit_len += en - st; } hit_len += feat_hit_len; if (print_len) print('F', t.slice(0, 4).join("\t"), feat_len, feat_hit_len); } file.close(); buf.destroy(); warn("# target bases: " + tot_len); warn("# target bases overlapping regions: " + hit_len + ' (' + (100.0 * hit_len / tot_len).toFixed(2) + '%)'); } /********************** * Conversion related * **********************/ function paf_view(args) { var c, line_len = 80, fmt = "aln"; while ((c = getopt(args, "f:l:")) != null) { if (c == 'f') { fmt = getopt.arg; if (fmt != "aln" && fmt != "lastz-cigar" && fmt != "maf") throw Error("format must be one of aln, lastz-cigar and maf"); } else if (c == 'l') line_len = parseInt(getopt.arg); } if (line_len == 0) line_len = 0x7fffffff; if (getopt.ind == args.length) { print("Usage: paftools.js view [options] "); print("Options:"); print(" -f STR output format: aln (BLAST-like), maf or lastz-cigar [aln]"); print(" -l INT line length in BLAST-like output [80]"); exit(1); } function padding_str(x, len, right) { var s = x.toString(); if (s.length < len) { if (right) s += Array(len - s.length + 1).join(" "); else s = Array(len - s.length + 1).join(" ") + s; } return s; } function update_aln(s_ref, s_qry, s_mid, type, seq, slen) { var l = type == '*'? 1 : seq.length; if (type == '=' || type == ':') { s_ref.set(seq); s_qry.set(seq); s_mid.set(Array(l+1).join("|")); slen[0] += l, slen[1] += l; } else if (type == '*') { s_ref.set(seq.charAt(0)); s_qry.set(seq.charAt(1)); s_mid.set(' '); slen[0] += 1, slen[1] += 1; } else if (type == '+') { s_ref.set(Array(l+1).join("-")); s_qry.set(seq); s_mid.set(Array(l+1).join(" ")); slen[1] += l; } else if (type == '-') { s_ref.set(seq); s_qry.set(Array(l+1).join("-")); s_mid.set(Array(l+1).join(" ")); slen[0] += l; } } function print_aln(rs, qs, strand, slen, elen, s_ref, s_qry, s_mid) { print(["Ref+:", padding_str(rs + slen[0] + 1, 10, false), s_ref.toString(), padding_str(rs + elen[0], 10, true)].join(" ")); print(" " + s_mid.toString()); var st, en; if (strand == '+') st = qs + slen[1] + 1, en = qs + elen[1]; else st = qs - slen[1], en = qs - elen[1] + 1; print(["Qry" + strand + ":", padding_str(st, 10, false), s_qry.toString(), padding_str(en, 10, true)].join(" ")); } var s_ref = new Bytes(), s_qry = new Bytes(), s_mid = new Bytes(); // these are used to show padded alignment var re_cs = /([:=\-\+\*])(\d+|[A-Za-z]+)/g; var re_cg = /(\d+)([MIDNSH])/g; var buf = new Bytes(); var file = args[getopt.ind] == "-"? new File() : new File(args[getopt.ind]); var lineno = 0; if (fmt == "maf") print("##maf version=1\n"); while (file.readline(buf) >= 0) { var m, line = buf.toString(); var t = line.split("\t", 12); ++lineno; s_ref.length = s_qry.length = s_mid.length = 0; var slen = [0, 0], elen = [0, 0]; if (fmt == "lastz-cigar") { // LASTZ-cigar output var cg = (m = /\tcg:Z:(\S+)/.exec(line)) != null? m[1] : null; if (cg == null) { warn("WARNING: converting to LASTZ-cigar format requires the 'cg' tag, which is absent on line " + lineno); continue; } var score = (m = /\tAS:i:(\d+)/.exec(line)) != null? m[1] : 0; var out = ['cigar:', t[0], t[2], t[3], t[4], t[5], t[7], t[8], '+', score]; while ((m = re_cg.exec(cg)) != null) out.push(m[2], m[1]); print(out.join(" ")); } else if (fmt == "maf") { // MAF output var cs = (m = /\tcs:Z:(\S+)/.exec(line)) != null? m[1] : null; if (cs == null) { warn("WARNING: converting to MAF requires the 'cs' tag, which is absent on line " + lineno); continue; } while ((m = re_cs.exec(cs)) != null) { if (m[1] == ':') throw Error("converting to MAF only works with 'minimap2 --cs=long'"); update_aln(s_ref, s_qry, s_mid, m[1], m[2], elen); } var score = (m = /\tAS:i:(\d+)/.exec(line)) != null? parseInt(m[1]) : 0; var len = t[0].length > t[5].length? t[0].length : t[5].length; print("a " + score); print(["s", padding_str(t[5], len, true), padding_str(t[7], 10, false), padding_str(parseInt(t[8]) - parseInt(t[7]), 10, false), "+", padding_str(t[6], 10, false), s_ref.toString()].join(" ")); var qs, qe, ql = parseInt(t[1]); if (t[4] == '+') { qs = parseInt(t[2]); qe = parseInt(t[3]); } else { qs = ql - parseInt(t[3]); qe = ql - parseInt(t[2]); } print(["s", padding_str(t[0], len, true), padding_str(qs, 10, false), padding_str(qe - qs, 10, false), t[4], padding_str(ql, 10, false), s_qry.toString()].join(" ")); print(""); } else { // BLAST-like output var cs = (m = /\tcs:Z:(\S+)/.exec(line)) != null? m[1] : null; if (cs == null) { warn("WARNING: converting to BLAST-like alignment requires the 'cs' tag, which is absent on line " + lineno); continue; } line = line.replace(/\tc[sg]:Z:\S+/g, ""); // get rid of cs or cg tags print('>' + line); var rs = parseInt(t[7]), qs = t[4] == '+'? parseInt(t[2]) : parseInt(t[3]); var n_blocks = 0; while ((m = re_cs.exec(cs)) != null) { if (m[1] == ':') m[2] = Array(parseInt(m[2]) + 1).join("="); var start = 0, rest = m[1] == '*'? 1 : m[2].length; while (rest > 0) { var l_proc; if (s_ref.length + rest >= line_len) { l_proc = line_len - s_ref.length; update_aln(s_ref, s_qry, s_mid, m[1], m[1] == '*'? m[2] : m[2].substr(start, l_proc), elen); if (n_blocks > 0) print(""); print_aln(rs, qs, t[4], slen, elen, s_ref, s_qry, s_mid); ++n_blocks; s_ref.length = s_qry.length = s_mid.length = 0; slen[0] = elen[0], slen[1] = elen[1]; } else { l_proc = rest; update_aln(s_ref, s_qry, s_mid, m[1], m[1] == '*'? m[2] : m[2].substr(start, l_proc), elen); } rest -= l_proc, start += l_proc; } } if (s_ref.length > 0) { if (n_blocks > 0) print(""); print_aln(rs, qs, t[4], slen, elen, s_ref, s_qry, s_mid); ++n_blocks; } print("//"); } } file.close(); buf.destroy(); s_ref.destroy(); s_qry.destroy(); s_mid.destroy(); } function paf_gff2bed(args) { var c, fn_ucsc_fai = null, is_short = false; while ((c = getopt(args, "u:s")) != null) { if (c == 'u') fn_ucsc_fai = getopt.arg; else if (c == 's') is_short = true; } if (getopt.ind == args.length) { print("Usage: paftools.js gff2bed [-u ucsc-genome.fa.fai] "); exit(1); } var ens2ucsc = {}; if (fn_ucsc_fai != null) { var buf = new Bytes(); var file = new File(fn_ucsc_fai); while (file.readline(buf) >= 0) { var t = buf.toString().split("\t"); var s = t[0]; if (/_(random|alt|decoy)$/.test(s)) { s = s.replace(/_(random|alt|decoy)$/, ''); s = s.replace(/^chr\S+_/, ''); } else { s = s.replace(/^chrUn_/, ''); } s = s.replace(/v(\d+)/, ".$1"); if (s != t[0]) ens2ucsc[s] = t[0]; } file.close(); buf.destroy(); } var colors = { 'protein_coding':'0,128,255', 'lincRNA':'0,192,0', 'snRNA':'0,192,0', 'miRNA':'0,192,0', 'misc_RNA':'0,192,0' }; function print_bed12(exons, cds_st, cds_en, is_short) { if (exons.length == 0) return; var name = is_short? exons[0][7] + "|" + exons[0][5] : exons[0].slice(4, 7).join("|"); var a = exons.sort(function(a,b) {return a[1]-b[1]}); var sizes = [], starts = [], st, en; st = a[0][1]; en = a[a.length - 1][2]; if (cds_st == 1<<30) cds_st = st; if (cds_en == 0) cds_en = en; if (cds_st < st || cds_en > en) throw Error("inconsistent thick start or end for transcript " + a[0][4]); for (var i = 0; i < a.length; ++i) { sizes.push(a[i][2] - a[i][1]); starts.push(a[i][1] - st); } var color = colors[a[0][5]]; if (color == null) color = '196,196,196'; print(a[0][0], st, en, name, 1000, a[0][3], cds_st, cds_en, color, a.length, sizes.join(",") + ",", starts.join(",") + ","); } var re_gtf = /(transcript_id|transcript_type|transcript_biotype|gene_name|transcript_name) "([^"]+)";/g; var re_gff3 = /(transcript_id|transcript_type|transcript_biotype|gene_name|transcript_name)=([^;]+)/g; var buf = new Bytes(); var file = args[getopt.ind] == '-'? new File() : new File(args[getopt.ind]); var exons = [], cds_st = 1<<30, cds_en = 0, last_id = null; while (file.readline(buf) >= 0) { var t = buf.toString().split("\t"); if (t[0].charAt(0) == '#') continue; if (t[2] != "CDS" && t[2] != "exon") continue; t[3] = parseInt(t[3]) - 1; t[4] = parseInt(t[4]); var id = null, type = "", gname = "N/A", biotype = "", m, tname = "N/A"; while ((m = re_gtf.exec(t[8])) != null) { if (m[1] == "transcript_id") id = m[2]; else if (m[1] == "transcript_type") type = m[2]; else if (m[1] == "transcript_biotype") biotype = m[2]; else if (m[1] == "gene_name") name = m[2]; else if (m[1] == "transcript_name") tname = m[2]; } while ((m = re_gff3.exec(t[8])) != null) { if (m[1] == "transcript_id") id = m[2]; else if (m[1] == "transcript_type") type = m[2]; else if (m[1] == "transcript_biotype") biotype = m[2]; else if (m[1] == "gene_name") name = m[2]; else if (m[1] == "transcript_name") tname = m[2]; } if (type == "" && biotype != "") type = biotype; if (id == null) throw Error("No transcript_id"); if (id != last_id) { print_bed12(exons, cds_st, cds_en, is_short); exons = [], cds_st = 1<<30, cds_en = 0; last_id = id; } if (t[2] == "CDS") { cds_st = cds_st < t[3]? cds_st : t[3]; cds_en = cds_en > t[4]? cds_en : t[4]; } else if (t[2] == "exon") { if (fn_ucsc_fai != null) { if (ens2ucsc[t[0]] != null) t[0] = ens2ucsc[t[0]]; else if (/^[A-Z]+\d+\.\d+$/.test(t[0])) t[0] = t[0].replace(/([A-Z]+\d+)\.(\d+)/, "chrUn_$1v$2"); } exons.push([t[0], t[3], t[4], t[6], id, type, name, tname]); } } if (last_id != null) print_bed12(exons, cds_st, cds_en, is_short); file.close(); buf.destroy(); } function paf_sam2paf(args) { var c, pri_only = false; while ((c = getopt(args, "p")) != null) if (c == 'p') pri_only = true; var file = args.length == getopt.ind || args[getopt.ind] == "-"? new File() : new File(args[getopt.ind]); var buf = new Bytes(); var re = /(\d+)([MIDSHNX=])/g; var len = {}, lineno = 0; while (file.readline(buf) >= 0) { var m, n_cigar = 0, line = buf.toString(); ++lineno; if (line.charAt(0) == '@') { if (/^@SQ/.test(line)) { var name = (m = /\tSN:(\S+)/.exec(line)) != null? m[1] : null; var l = (m = /\tLN:(\d+)/.exec(line)) != null? parseInt(m[1]) : null; if (name != null && l != null) len[name] = l; } continue; } var t = line.split("\t"); var flag = parseInt(t[1]); if (t[9] != '*' && t[10] != '*' && t[9].length != t[10].length) throw Error("ERROR at line " + lineno + ": inconsistent SEQ and QUAL lengths - " + t[9].length + " != " + t[10].length); if (t[2] == '*' || (flag&4)) continue; if (pri_only && (flag&0x100)) continue; var tlen = len[t[2]]; if (tlen == null) throw Error("ERROR at line " + lineno + ": can't find the length of contig " + t[2]); var nn = (m = /\tnn:i:(\d+)/.exec(line)) != null? parseInt(m[1]) : 0; var NM = (m = /\tNM:i:(\d+)/.exec(line)) != null? parseInt(m[1]) : null; var have_NM = NM == null? false : true; NM += nn; var clip = [0, 0], I = [0, 0], D = [0, 0], M = 0, N = 0, ql = 0, tl = 0, mm = 0, ext_cigar = false; while ((m = re.exec(t[5])) != null) { var l = parseInt(m[1]); if (m[2] == 'M') M += l, ql += l, tl += l, ext_cigar = false; else if (m[2] == 'I') ++I[0], I[1] += l, ql += l; else if (m[2] == 'D') ++D[0], D[1] += l, tl += l; else if (m[2] == 'N') N += l, tl += l; else if (m[2] == 'S') clip[M == 0? 0 : 1] = l, ql += l; else if (m[2] == 'H') clip[M == 0? 0 : 1] = l; else if (m[2] == '=') M += l, ql += l, tl += l, ext_cigar = true; else if (m[2] == 'X') M += l, ql += l, tl += l, mm += l, ext_cigar = true; ++n_cigar; } if (n_cigar > 65535) warn("WARNING at line " + lineno + ": " + n_cigar + " CIGAR operations"); if (tl + parseInt(t[3]) - 1 > tlen) { warn("WARNING at line " + lineno + ": alignment end position larger than ref length; skipped"); continue; } if (t[9] != '*' && t[9].length != ql) { warn("WARNING at line " + lineno + ": SEQ length inconsistent with CIGAR (" + t[9].length + " != " + ql + "); skipped"); continue; } if (!have_NM || ext_cigar) NM = I[1] + D[1] + mm; if (NM < I[1] + D[1] + mm) { warn("WARNING at line " + lineno + ": NM is less than the total number of gaps (" + NM + " < " + (I[1]+D[1]+mm) + ")"); NM = I[1] + D[1] + mm; } var extra = ["mm:i:"+(NM-I[1]-D[1]), "io:i:"+I[0], "in:i:"+I[1], "do:i:"+D[0], "dn:i:"+D[1]]; var match = M - (NM - I[1] - D[1]); var blen = M + I[1] + D[1]; var qlen = M + I[1] + clip[0] + clip[1]; var qs, qe; if (flag&16) qs = clip[1], qe = qlen - clip[0]; else qs = clip[0], qe = qlen - clip[1]; var ts = parseInt(t[3]) - 1, te = ts + M + D[1] + N; var qname = t[0]; if ((flag&1) && (flag&0x40)) qname += '/1'; if ((flag&1) && (flag&0x80)) qname += '/2'; var a = [qname, qlen, qs, qe, flag&16? '-' : '+', t[2], tlen, ts, te, match, blen, t[4]]; print(a.join("\t"), extra.join("\t")); } buf.destroy(); file.close(); } function paf_delta2paf(args) { if (args.length == 0) { print("Usage: paftools.js delta2paf "); exit(1); } var buf = new Bytes(); var file = args[0] == '-'? new File() : new File(args[0]); var rname, qname, rlen, qlen, qs, qe, rs, re, strand, NM, cigar, x, y, seen_gt = false; while (file.readline(buf) >= 0) { var m, line = buf.toString(); if ((m = /^>(\S+)\s+(\S+)\s+(\d+)\s+(\d+)/.exec(line)) != null) { rname = m[1], qname = m[2], rlen = parseInt(m[3]), qlen = parseInt(m[4]); seen_gt = true; continue; } if (!seen_gt) continue; var t = line.split(" "); if (t.length == 7) { for (var i = 0; i < 5; ++i) t[i] = parseInt(t[i]); strand = ((t[0] < t[1] && t[2] < t[3]) || (t[0] > t[1] && t[2] > t[3]))? 1 : -1; rs = (t[0] < t[1]? t[0] : t[1]) - 1; re = t[1] > t[0]? t[1] : t[0]; qs = (t[2] < t[3]? t[2] : t[3]) - 1; qe = t[3] > t[2]? t[3] : t[2]; x = y = 0; NM = parseInt(t[4]); cigar = []; } else if (t.length == 1) { var d = parseInt(t[0]); if (d == 0) { var blen = 0, cigar_str = []; if (re - rs - x != qe - qs - y) throw Error("inconsisnt alignment"); cigar.push((re - rs - x) << 4); for (var i = 0; i < cigar.length; ++i) { blen += cigar[i] >> 4; cigar_str.push((cigar[i]>>4) + "MID".charAt(cigar[i]&0xf)); } print([qname, qlen, qs, qe, strand > 0? '+' : '-', rname, rlen, rs, re, blen - NM, blen, 0, "NM:i:" + NM, "cg:Z:" + cigar_str.join("")].join("\t")); } else if (d > 0) { var l = d - 1; x += l + 1, y += l; if (l > 0) cigar.push(l<<4); if (cigar.length > 0 && (cigar[cigar.length-1]&0xf) == 2) cigar[cigar.length-1] += 1<<4; else cigar.push(1<<4|2); // deletion } else { var l = -d - 1; x += l, y += l + 1; if (l > 0) cigar.push(l<<4); if (cigar.length > 0 && (cigar[cigar.length-1]&0xf) == 1) cigar[cigar.length-1] += 1<<4; else cigar.push(1<<4|1); // insertion } } } file.close(); buf.destroy(); } function paf_splice2bed(args) { var colors = ["0,128,255", "255,0,0", "0,192,0"]; function print_lines(a, fmt, keep_multi) { if (a.length == 0) return; if (fmt == "bed") { var n_pri = 0; for (var i = 0; i < a.length; ++i) if (a[i][8] == 0) ++n_pri; if (n_pri > 1) { for (var i = 0; i < a.length; ++i) if (a[i][8] == 0) a[i][8] = 1; } else if (n_pri == 0) { warn("Warning: " + a[0][3] + " doesn't have a primary alignment"); } for (var i = 0; i < a.length; ++i) { if (!keep_multi && a[i][8] == 2) continue; a[i][8] = colors[a[i][8]]; print(a[i].join("\t")); } } a.length = 0; } var re = /(\d+)([MIDNSH])/g; var c, fmt = "bed", fn_name_conv = null, keep_multi = false; while ((c = getopt(args, "f:n:m")) != null) { if (c == 'f') fmt = getopt.arg; else if (c == 'n') fn_name_conv = getopt.arg; else if (c == 'm') keep_multi = true; } if (getopt.ind == args.length) { print("Usage: paftools.js splice2bed [options] |"); print("Options:"); print(" -m keep multiple mappings (SAM flag 0x100)"); exit(1); } var conv = null; if (fn_name_conv != null) { conv = new Map(); var file = new File(fn_name_conv); var buf = new Bytes(); while (file.readline(buf) >= 0) { var t = buf.toString().split("\t"); conv.put(t[0], t[1]); } buf.destroy(); file.close(); } var file = args[getopt.ind] == '-'? new File() : new File(args[getopt.ind]); var buf = new Bytes(); var a = []; while (file.readline(buf) >= 0) { var line = buf.toString(); if (line.charAt(0) == '@') continue; // skip SAM header lines var t = line.split("\t"); var is_pri = false, cigar = null, a1; var qname = conv != null? conv.get(t[0]) : null; if (qname != null) t[0] = qname; if (t.length >= 10 && t[4] != '+' && t[4] != '-' && /^\d+/.test(t[1])) { // SAM var flag = parseInt(t[1]); if (flag&1) t[0] += '/' + (flag>>6&3); } if (a.length && a[0][3] != t[0]) { print_lines(a, fmt, keep_multi); a = []; } if (t.length >= 12 && (t[4] == '+' || t[4] == '-')) { // PAF for (var i = 12; i < t.length; ++i) { if (t[i].substr(0, 5) == 'cg:Z:') { cigar = t[i].substr(5); } else if (t[i].substr(0, 5) == 's2:i:') { is_pri = true; } } a1 = [t[5], t[7], t[8], t[0], Math.floor(t[9]/t[10]*1000), t[4]]; } else if (t.length >= 10) { // SAM var flag = parseInt(t[1]); if ((flag&4) || a[2] == '*') continue; cigar = t[5]; is_pri = (flag&0x100)? false : true; a1 = [t[2], parseInt(t[3])-1, null, t[0], 1000, (flag&16)? '-' : '+']; } else { throw Error("unrecognized input format"); } if (cigar == null) throw Error("missing CIGAR"); var m, x0 = 0, x = 0, bs = [], bl = []; while ((m = re.exec(cigar)) != null) { if (m[2] == 'M' || m[2] == 'D') { x += parseInt(m[1]); } else if (m[2] == 'N') { bs.push(x0); bl.push(x - x0); x += parseInt(m[1]); x0 = x; } } bs.push(x0); bl.push(x - x0); // write the BED12 line if (a1[2] == null) a1[2] = a1[1] + x; a1.push(a1[1], a1[2]); // thick start/end is the same as start/end a1.push(is_pri? 0 : 2, bs.length, bl.join(",")+",", bs.join(",")+","); a.push(a1); } print_lines(a, fmt, keep_multi); buf.destroy(); file.close(); if (conv != null) conv.destroy(); } /********************** * Evaluation related * **********************/ // evaluate mapping accuracy function paf_mapeval(args) { var c, max_mapq = 60, mode = 0, err_out_q = 256, print_err = false, ovlp_ratio = 0.1, cap_short_mapq = false; while ((c = getopt(args, "Q:r:m:c")) != null) { if (c == 'Q') err_out_q = parseInt(getopt.arg), print_err = true; else if (c == 'r') ovlp_ratio = parseFloat(getopt.arg); else if (c == 'm') mode = parseInt(getopt.arg); else if (c == 'c') cap_short_mapq = true; } if (args.length == getopt.ind) { warn("Usage: paftools.js mapeval [options] |"); warn("Options:"); warn(" -r FLOAT mapping correct if overlap_length/union_length>FLOAT [" + ovlp_ratio + "]"); warn(" -Q INT print wrong mappings with mapQ>INT [don't print]"); warn(" -m INT 0: eval the longest aln only; 1: first aln only; 2: all primary aln [0]"); exit(1); } var file = args[getopt.ind] == '-'? new File() : new File(args[getopt.ind]); var buf = new Bytes(); var tot = [], err = []; for (var q = 0; q <= max_mapq; ++q) tot[q] = err[q] = 0; function is_correct(s, b) { if (s[0] != b[0] || s[3] != b[3]) return false; var o, l; if (s[1] < b[1]) { if (s[2] <= b[1]) return false; o = (s[2] < b[2]? s[2] : b[2]) - b[1]; l = (s[2] > b[2]? s[2] : b[2]) - s[1]; } else { if (b[2] <= s[1]) return false; o = (s[2] < b[2]? s[2] : b[2]) - s[1]; l = (s[2] > b[2]? s[2] : b[2]) - b[1]; } return o/l > ovlp_ratio? true : false; } function count_err(qname, a, tot, err, mode) { if (a.length == 0) return; var m, s; if ((m = /^(\S+)!(\S+)!(\d+)!(\d+)!([\+\-])$/.exec(qname)) != null) { // pbsim single-end reads s = [m[1], m[2], parseInt(m[3]), parseInt(m[4]), m[5]]; } else if ((m = /^(\S+)!(\S+)!(\d+)_(\d+)!(\d+)_(\d+)!([\+\-])([\+\-])\/([12])$/.exec(qname)) != null) { // mason2 paired-end reads if (m[9] == '1') { s = [m[1], m[2], parseInt(m[3]), parseInt(m[5]), m[7]]; } else { s = [m[1], m[2], parseInt(m[4]), parseInt(m[6]), m[8]]; } } else throw Error("Failed to parse simulated read names '" + qname + "'"); s.shift(); // skip the orginal read name if (mode == 0 || mode == 1) { // longest only or first only var max_i = 0; if (mode == 0) { // longest only var max = 0; for (var i = 0; i < a.length; ++i) if (a[i][5] > max) max = a[i][5], max_i = i; } var mapq = a[max_i][4]; ++tot[mapq]; if (!is_correct(s, a[max_i])) { if (mapq >= err_out_q) print('E', qname, a[max_i].join("\t")); ++err[mapq]; } } else if (mode == 2) { // all primary mode var max_err_mapq = -1, max_mapq = 0, max_err_i = -1; if (cap_short_mapq) { var max = 0, max_q = 0; for (var i = 0; i < a.length; ++i) if (a[i][5] > max) max = a[i][5], max_q = a[i][4]; for (var i = 0; i < a.length; ++i) a[i][4] = max_q < a[i][4]? max_q : a[i][4]; } for (var i = 0; i < a.length; ++i) { max_mapq = max_mapq > a[i][4]? max_mapq : a[i][4]; if (!is_correct(s, a[i])) if (a[i][4] > max_err_mapq) max_err_mapq = a[i][4], max_err_i = i; } if (max_err_mapq >= 0) { ++tot[max_err_mapq], ++err[max_err_mapq]; if (max_err_mapq >= err_out_q) print('E', qname, a[max_err_i].join("\t")); } else ++tot[max_mapq]; } } var lineno = 0, last = null, a = [], n_unmapped = null; var re_cigar = /(\d+)([MIDSHN])/g; while (file.readline(buf) >= 0) { var m, line = buf.toString(); ++lineno; if (line[0] != '@') { var t = line.split("\t"); if (t[4] == '+' || t[4] == '-') { // PAF if (last != t[0]) { if (last != null) count_err(last, a, tot, err, mode); a = [], last = t[0]; } if (/\ts1:i:\d+/.test(line) && !/\ts2:i:\d+/.test(line)) // secondary alignment in minimap2 PAF continue; var mapq = parseInt(t[11]); if (mapq > max_mapq) mapq = max_mapq; a.push([t[5], parseInt(t[7]), parseInt(t[8]), t[4], mapq, parseInt(t[9])]); } else { // SAM var flag = parseInt(t[1]); var read_no = flag>>6&0x3; var qname = t[0]; if (!/\/[12]$/.test(qname)) qname = read_no == 1 || read_no == 2? t[0] + '/' + read_no : t[0]; if (last != qname) { if (last != null) count_err(last, a, tot, err, mode); a = [], last = qname; } if (flag&0x100) continue; // secondary alignment if ((flag&0x4) || t[2] == '*') { // unmapped if (n_unmapped == null) n_unmapped = 0; ++n_unmapped; continue; } var mapq = parseInt(t[4]); if (mapq > max_mapq) mapq = max_mapq; var pos = parseInt(t[3]) - 1, pos_end = pos; var n_gap = 0, mlen = 0; while ((m = re_cigar.exec(t[5])) != null) { var len = parseInt(m[1]); if (m[2] == 'M') pos_end += len, mlen += len; else if (m[2] == 'I') n_gap += len; else if (m[2] == 'D') n_gap += len, pos_end += len; } var score = pos_end - pos; if ((m = /\tNM:i:(\d+)/.exec(line)) != null) { var NM = parseInt(m[1]); if (NM >= n_gap) score = mlen - (NM - n_gap); } a.push([t[2], pos, pos_end, (flag&16)? '-' : '+', mapq, score]); } } } if (last != null) count_err(last, a, tot, err, mode); buf.destroy(); file.close(); var sum_tot = 0, sum_err = 0, q_out = -1, sum_tot2 = 0, sum_err2 = 0; for (var q = max_mapq; q >= 0; --q) { if (tot[q] == 0) continue; if (q_out < 0 || err[q] > 0) { if (q_out >= 0) print('Q', q_out, sum_tot, sum_err, (sum_err2/sum_tot2).toFixed(9), sum_tot2); sum_tot = sum_err = 0, q_out = q; } sum_tot += tot[q], sum_err += err[q]; sum_tot2 += tot[q], sum_err2 += err[q]; } print('Q', q_out, sum_tot, sum_err, (sum_err2/sum_tot2).toFixed(9), sum_tot2); if (n_unmapped != null) print('U', n_unmapped); } // convert mason2 SAM to FASTQ function paf_mason2fq(args) { if (args.length == 0) { print("Usage: paftools.js mason2fq "); exit(1); } function print_se(a) { print('@' + a.slice(0, 5).join("!") + " " + a[8]); print(a[5]); print("+"); print(a[6]); } var buf = new Bytes(), buf2 = new Bytes(); var file = new File(args[0]); var re = /(\d+)([MIDSHN])/g; var last = null; while (file.readline(buf) >= 0) { var t = buf.toString().split("\t"); if (t[0].charAt(0) == '@') continue; var m, l_ref = 0; while ((m = re.exec(t[5])) != null) if (m[2] == 'D' || m[2] == 'M' || m[2] == 'N') l_ref += parseInt(m[1]); var flag = parseInt(t[1]); var rev = !!(flag&16); var seq, qual; if (rev) { buf2.length = 0; buf2.set(t[9], 0); buf2.revcomp(); seq = buf2.toString(); buf2.set(t[10], 0); buf2.reverse(); qual = buf2.toString(); } else seq = t[9], qual = t[10]; var qname = t[0]; qname = qname.replace(/^simulated./, ""); var chr = t[2]; var pos = parseInt(t[3]) - 1; var strand = (flag&16)? '-' : '+'; var read_no = flag&0xc0; if (read_no == 0x40) read_no = 1; else if (read_no == 0x80) read_no = 2; else read_no = 0; var err = 0, snp = 0, indel = 0; for (var i = 11; i < t.length; ++i) { if ((m = /^XE:i:(\d+)/.exec(t[i])) != null) err = m[1]; else if ((m = /^XS:i:(\d+)/.exec(t[i])) != null) snp = m[1]; else if ((m = /^XI:i:(\d+)/.exec(t[i])) != null) indel = m[1]; } var comment = [err, snp, indel].join(":"); if (last == null) { last = [qname, chr, pos, pos + l_ref, strand, seq, qual, read_no, comment]; } else if (last[0] != qname) { print_se(last); last = [qname, chr, pos, pos + l_ref, strand, seq, qual, read_no, comment]; } else { if (read_no == 2) { // last[] is the first read if (last[7] != 1) throw Error("ERROR: can't find read1"); var name = [qname, chr, last[2] + "_" + pos, last[3] + "_" + (pos + l_ref), last[4] + strand].join("!"); print('@' + name + '/1' + ' ' + last[8]); print(last[5]); print("+"); print(last[6]); print('@' + name + '/2' + ' ' + comment); print(seq); print("+"); print(qual); } else { if (last[7] != 2) throw Error("ERROR: can't find read2"); var name = [qname, chr, pos + "_" + last[2], (pos + l_ref) + "_" + last[3], strand + last[4]].join("!"); print('@' + name + '/1' + ' ' + comment); print(seq); print("+"); print(qual); print('@' + name + '/2' + ' ' + last[8]); print(last[5]); print("+"); print(last[6]); } last = null; } } if (last != null) print_se(last); file.close(); buf.destroy(); buf2.destroy(); } // convert pbsim MAF to FASTQ function paf_pbsim2fq(args) { if (args.length < 2) { print("Usage: paftools.js pbsim2fq [[pbsim2.maf] ...]"); exit(1); } var file, buf = new Bytes(), buf2 = new Bytes(); file = new File(args[0]); var chr_list = []; while (file.readline(buf) >= 0) { var t = buf.toString().split(/\s+/); chr_list.push(t[0]); } file.close(); for (var k = 1; k < args.length; ++k) { var fn = args[k]; file = new File(fn); var state = 0, reg; while (file.readline(buf) >= 0) { var line = buf.toString(); if (state == 0 && line.charAt(0) == 'a') { state = 1; } else if (state == 1 && line.charAt(0) == 's') { var t = line.split(/\s+/); var st = parseInt(t[2]); reg = [st, st + parseInt(t[3])]; state = 2; } else if (state == 2 && line.charAt(0) == 's') { var m, t = line.split(/\s+/); if ((m = /S(\d+)_\d+/.exec(t[1])) == null) throw Error("Failed to parse the read name"); var chr_id = parseInt(m[1]) - 1; if (chr_id >= chr_list.length) throw Error("Index outside the chr list"); var name = [t[1], chr_list[chr_id], reg[0], reg[1], t[4]].join("!"); var seq = t[6].replace(/\-/g, ""); if (seq.length != parseInt(t[5])) throw Error("Inconsistent read length"); if (seq.indexOf("NN") < 0) { if (t[4] == '-') { buf2.set(seq, 0); buf2.length = seq.length; buf2.revcomp(); seq = buf2.toString(); } print(">" + name); print(seq); } state = 0; } } file.close(); } buf.destroy(); buf2.destroy(); } function paf_junceval(args) { var c, l_fuzzy = 0, print_ovlp = false, print_err_only = false, first_only = false; while ((c = getopt(args, "l:ep")) != null) { if (c == 'l') l_fuzzy = parseInt(getopt.arg); else if (c == 'e') print_err_only = print_ovlp = true; else if (c == 'p') print_ovlp = true; } if (args.length - getopt.ind < 2) { print("Usage: paftools.js junceval [options] "); print("Options:"); print(" -l INT tolerance of junction positions (0 for exact) [0]"); print(" -p print overlapping introns"); print(" -e print erroreous overlapping introns"); exit(1); } var file, buf = new Bytes(); var tr = {}; file = new File(args[getopt.ind]); while (file.readline(buf) >= 0) { var m, t = buf.toString().split("\t"); if (t[0].charAt(0) == '#') continue; if (t[2] != 'exon') continue; var st = parseInt(t[3]) - 1; var en = parseInt(t[4]); if ((m = /transcript_id "(\S+)"/.exec(t[8])) == null) continue; var tid = m[1]; if (tr[tid] == null) tr[tid] = [t[0], t[6], 0, 0, []]; tr[tid][4].push([st, en]); } file.close(); var anno = {}; for (var tid in tr) { var t = tr[tid]; Interval.sort(t[4]); t[2] = t[4][0][0]; t[3] = t[4][t[4].length - 1][1]; if (anno[t[0]] == null) anno[t[0]] = []; var s = t[4]; for (var i = 0; i < s.length - 1; ++i) { if (s[i][1] >= s[i+1][0]) warn("WARNING: incorrect annotation for transcript "+tid+" ("+s[i][1]+" >= "+s[i+1][0]+")") anno[t[0]].push([s[i][1], s[i+1][0]]); } } tr = null; for (var chr in anno) { var e = anno[chr]; if (e.length == 0) continue; Interval.sort(e); var k = 0; for (var i = 1; i < e.length; ++i) // dedup if (e[i][0] != e[k][0] || e[i][1] != e[k][1]) e[++k] = e[i].slice(0); e.length = k + 1; Interval.index_end(e); } var n_pri = 0, n_unmapped = 0, n_mapped = 0; var n_sgl = 0, n_splice = 0, n_splice_hit = 0, n_splice_novel = 0; file = new File(args[getopt.ind+1]); var last_qname = null; var re_cigar = /(\d+)([MIDNSHX=])/g; while (file.readline(buf) >= 0) { var m, t = buf.toString().split("\t"); if (t[0].charAt(0) == '@') continue; var flag = parseInt(t[1]); if (flag&0x100) continue; if (first_only && last_qname == t[0]) continue; if (t[2] == '*') { ++n_unmapped; continue; } else { ++n_pri; if (last_qname != t[0]) { ++n_mapped; last_qname = t[0]; } } var pos = parseInt(t[3]) - 1, intron = []; while ((m = re_cigar.exec(t[5])) != null) { var len = parseInt(m[1]), op = m[2]; if (op == 'N') { intron.push([pos, pos + len]); pos += len; } else if (op == 'M' || op == 'X' || op == '=' || op == 'D') pos += len; } if (intron.length == 0) { ++n_sgl; continue; } n_splice += intron.length; var chr = anno[t[2]]; if (chr != null) { for (var i = 0; i < intron.length; ++i) { var o = Interval.find_ovlp(chr, intron[i][0], intron[i][1]); if (o.length > 0) { var hit = false; for (var j = 0; j < o.length; ++j) { var st_diff = intron[i][0] - o[j][0]; var en_diff = intron[i][1] - o[j][1]; if (st_diff < 0) st_diff = -st_diff; if (en_diff < 0) en_diff = -en_diff; if (st_diff <= l_fuzzy && en_diff <= l_fuzzy) ++n_splice_hit, hit = true; if (hit) break; } if (print_ovlp) { var type = hit? 'C' : 'P'; if (hit && print_err_only) continue; var x = '['; for (var j = 0; j < o.length; ++j) { if (j) x += ', '; x += '(' + o[j][0] + "," + o[j][1] + ')'; } x += ']'; print(type, t[0], i+1, t[2], intron[i][0], intron[i][1], x); } } else { ++n_splice_novel; if (print_ovlp) print('N', t[0], i+1, t[2], intron[i][0], intron[i][1]); } } } else { n_splice_novel += intron.length; } } file.close(); buf.destroy(); if (!print_ovlp) { print("# unmapped reads: " + n_unmapped); print("# mapped reads: " + n_mapped); print("# primary alignments: " + n_pri); print("# singletons: " + n_sgl); print("# predicted introns: " + n_splice); print("# non-overlapping introns: " + n_splice_novel); print("# correct introns: " + n_splice_hit + " (" + (n_splice_hit / n_splice * 100).toFixed(2) + "%)"); } } // evaluate overlap sensitivity function paf_ov_eval(args) { var c, min_ovlp = 2000, min_frac = 0.95, min_mapq = 10; while ((c = getopt(args, "q:l:f:")) != null) { if (c == 'q') min_mapq = parseInt(getopt.arg); else if (c == 'l') min_ovlp = parseInt(getopt.arg); else if (c == 'f') min_frac = parseFloat(getopt.arg); } if (args.length - getopt.ind < 2) { print("Usage: sort -k6,6 -k8,8n to-ref.paf | paftools.js ov-eval [options] - "); print("Options:"); print(" -l INT min overlap length [2000]"); print(" -q INT min mapping quality [10]"); print(" -f FLOAT min fraction of mapped length [0.95]"); exit(1); } var buf = new Bytes(); var file = args[getopt.ind] == '-'? new File() : new File(args[getopt.ind]); var a = [], h = {}; while (file.readline(buf) >= 0) { var t = buf.toString().split("\t"); var is_pri = false; if (parseInt(t[11]) < min_mapq) continue; for (var i = 12; i < t.length; ++i) if (t[i] == 'tp:A:P') is_pri = true; if (!is_pri) continue; for (var i = 1; i <= 3; ++i) t[i] = parseInt(t[i]); for (var i = 6; i <= 8; ++i) t[i] = parseInt(t[i]); if (t[3] - t[2] < min_ovlp || t[8] - t[7] < min_ovlp || (t[3] - t[2]) / t[1] < min_frac) continue; var ctg = t[5], st = t[7], en = t[8]; while (a.length > 0) { if (a[0][0] == ctg && a[0][2] > st) break; else a.shift(); } for (var j = 0; j < a.length; ++j) { if (a[j][3] == t[0]) continue; var len = (en > a[j][2]? a[j][2] : en) - st; if (len >= min_ovlp) { var key = a[j][3] < t[0]? a[j][3] + "\t" + t[0] : t[0] + "\t" + a[j][3]; h[key] = len; } } a.push([ctg, st, en, t[0]]); } file.close(); file = new File(args[getopt.ind + 1]); while (file.readline(buf) >= 0) { var t = buf.toString().split("\t"); var key = t[0] < t[5]? t[0] + "\t" + t[5] : t[5] + "\t" + t[0]; if (h[key] > 0) h[key] = -h[key]; } file.close(); buf.destroy(); var n_ovlp = 0, n_missing = 0; for (var key in h) { ++n_ovlp; if (h[key] > 0) ++n_missing; } print(n_ovlp + " overlaps inferred from the reference mapping"); print(n_missing + " missed by the read overlapper"); print((100 * (1 - n_missing / n_ovlp)).toFixed(2) + "% sensitivity"); } /************************* ***** main function ***** *************************/ function main(args) { if (args.length == 0) { print("Usage: paftools.js [arguments]"); print("Commands:"); print(" view convert PAF to BLAST-like (for eyeballing) or MAF"); print(" splice2bed convert spliced alignment in PAF/SAM to BED12"); print(" sam2paf convert SAM to PAF"); print(" delta2paf convert MUMmer's delta to PAF"); print(" gff2bed convert GTF/GFF3 to BED12"); print(""); print(" stat collect basic mapping information in PAF/SAM"); print(" liftover simplistic liftOver"); print(" call call variants from asm-to-ref alignment with the cs tag"); print(" bedcov compute the number of bases covered"); print(" version print paftools.js version"); print(""); print(" mapeval evaluate mapping accuracy using mason2/PBSIM-simulated FASTQ"); print(" mason2fq convert mason2-simulated SAM to FASTQ"); print(" pbsim2fq convert PBSIM-simulated MAF to FASTQ"); print(" junceval evaluate splice junction consistency with known annotations"); print(" ov-eval evaluate read overlap sensitivity using read-to-ref mapping"); exit(1); } var cmd = args.shift(); if (cmd == 'view') paf_view(args); else if (cmd == 'sam2paf') paf_sam2paf(args); else if (cmd == 'delta2paf') paf_delta2paf(args); else if (cmd == 'splice2bed') paf_splice2bed(args); else if (cmd == 'gff2bed') paf_gff2bed(args); else if (cmd == 'stat') paf_stat(args); else if (cmd == 'liftover' || cmd == 'liftOver') paf_liftover(args); else if (cmd == 'call') paf_call(args); else if (cmd == 'mapeval') paf_mapeval(args); else if (cmd == 'bedcov') paf_bedcov(args); else if (cmd == 'mason2fq') paf_mason2fq(args); else if (cmd == 'pbsim2fq') paf_pbsim2fq(args); else if (cmd == 'junceval') paf_junceval(args); else if (cmd == 'ov-eval') paf_ov_eval(args); else if (cmd == 'version') print(paftools_version); else throw Error("unrecognized command: " + cmd); } main(arguments);