1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60 61 62 63 64 65 66 67 68 69 70 71 72 73 74 75 76 77 78 79 80 81 82 83 84 85 86 87 88 89 90 91 92 93 94 95 96 97 98 99 100 101 102 103 104 105 106 107 108 109 110 111 112 113 114 115 116 117 118 119 120 121 122 123 124 125 126 127 128 129 130 131 132 133 134 135 136 137 138 139 140 141 142 143 144 145 146 147 148 149 150 151 152 153 154 155 156 157 158 159 160 161 162 163 164 165 166 167 168 169 170 171 172 173 174 175 176 177 178 179 180 181 182 183 184 185 186 187 188 189 190 191 192 193 194 195 196 197 198 199 200 201 202 203 204 205 206 207 208 209 210 211 212 213 214 215 216 217 218 219 220 221 222 223 224 225 226 227 228 229 230 231 232 233 234 235 236 237 238 239 240 241 242 243 244 245 246 247 248 249 250 251 252 253 254 255 256 257 258 259 260 261 262 263 264 265 266 267 268 269 270 271 272 273 274 275 276 277 278 279 280 281 282 283 284 285 286 287 288 289 290 291 292 293 294 295 296 297 298 299 300 301 302 303 304 305 306 307 308 309 310 311 312 313 314 315 316 317 318 319 320 321 322 323 324 325 326 327 328 329 330 331 332 333 334 335 336 337 338 339 340 341 342 343 344 345 346 347 348 349 350 351 352 353 354 355 356 357 358 359 360 361 362 363 364 365 366 367 368 369 370 371 372 373 374 375 376 377 378 379 380 381 382 383 384 385 386 387 388 389 390 391 392 393 394 395 396 397 398 399 400 401 402 403 404 405 406 407 408 409 410 411 412 413 414 415 416 417 418 419 420 421 422 423 424 425 426 427 428 429 430 431 432 433 434 435 436 437 438 439 440 441 442 443 444 445 446 447 448 449 450 451 452 453 454 455 456 457 458 459 460 461 462 463 464 465 466 467 468 469 470 471 472 473 474 475 476 477 478 479 480 481 482 483 484 485 486 487 488 489 490 491 492 493 494 495 496 497 498 499 500 501 502 503 504 505 506 507 508 509 510 511 512 513 514 515 516 517 518 519 520 521 522 523 524 525 526 527 528 529 530 531 532 533 534 535 536 537 538 539 540 541 542 543 544 545 546 547 548 549 550 551 552 553 554 555 556 557 558 559 560 561 562 563 564 565 566 567 568 569 570 571 572 573 574 575 576 577 578 579 580 581 582 583 584 585 586 587 588 589 590 591 592 593 594 595 596 597 598 599 600 601 602 603 604 605 606 607 608 609 610 611 612 613 614 615 616 617 618 619 620 621 622 623 624 625 626 627 628 629 630 631 632 633 634 635 636 637 638 639 640 641 642 643 644 645 646 647 648 649 650 651 652 653 654 655 656 657 658 659 660 661 662 663 664 665 666 667 668 669 670 671 672 673 674 675 676 677 678 679 680 681 682 683 684 685 686 687 688 689 690 691 692 693 694 695 696 697 698 699 700 701 702 703 704 705 706 707 708 709 710 711 712 713 714 715 716 717 718 719 720 721 722 723 724 725 726 727 728 729 730 731 732 733 734 735 736 737 738 739 740 741 742 743 744 745 746 747 748 749 750 751 752 753 754 755 756 757 758 759 760 761 762 763 764 765 766 767 768 769 770 771 772 773 774 775 776 777 778 779 780 781 782 783 784 785 786 787 788 789 790 791 792 793 794 795 796 797 798 799 800 801 802 803 804 805 806 807 808 809 810 811 812 813 814 815 816 817 818 819 820 821 822 823 824 825 826 827 828 829 830 831 832 833 834 835 836 837 838 839 840 841 842 843 844 845 846 847 848 849 850 851 852 853 854 855 856 857 858 859 860 861 862 863 864 865 866 867 868 869 870 871 872 873 874 875 876 877 878 879 880 881 882 883 884 885 886 887 888 889 890 891 892 893 894 895 896 897 898 899 900 901 902 903 904 905 906 907 908 909 910 911 912 913 914 915 916 917 918 919 920 921 922 923 924 925 926 927 928 929 930 931 932 933 934 935 936 937 938 939 940 941 942 943 944 945 946 947 948 949 950 951 952 953 954 955 956 957 958 959 960 961 962 963 964 965 966 967 968 969 970 971 972 973 974 975 976 977 978 979 980 981 982 983 984 985 986 987 988 989 990 991 992 993 994 995 996 997 998 999 1000 1001 1002 1003 1004 1005 1006 1007 1008 1009 1010 1011 1012 1013 1014 1015 1016 1017 1018 1019 1020 1021 1022 1023 1024 1025 1026 1027 1028 1029 1030 1031 1032 1033 1034 1035 1036 1037 1038 1039 1040 1041 1042 1043 1044 1045 1046 1047 1048 1049 1050 1051 1052 1053 1054 1055 1056 1057 1058 1059 1060 1061 1062 1063 1064 1065 1066 1067 1068 1069 1070 1071 1072 1073 1074 1075 1076 1077 1078 1079 1080 1081 1082 1083 1084 1085 1086 1087 1088 1089 1090 1091 1092 1093 1094 1095 1096 1097 1098 1099 1100 1101 1102 1103 1104 1105 1106 1107 1108 1109 1110 1111 1112 1113 1114 1115 1116 1117 1118 1119 1120 1121 1122 1123 1124 1125 1126 1127 1128 1129 1130 1131 1132 1133 1134 1135 1136 1137 1138 1139 1140 1141 1142 1143 1144 1145 1146 1147 1148 1149 1150 1151 1152 1153 1154 1155 1156 1157 1158 1159 1160 1161 1162 1163 1164 1165 1166 1167 1168 1169 1170 1171 1172 1173 1174 1175 1176 1177 1178 1179 1180 1181 1182 1183 1184 1185 1186 1187 1188 1189 1190 1191 1192 1193 1194 1195 1196 1197 1198 1199 1200 1201 1202 1203 1204 1205 1206 1207 1208 1209 1210 1211 1212 1213 1214 1215 1216 1217 1218 1219 1220 1221 1222 1223 1224 1225 1226 1227 1228 1229 1230 1231 1232 1233 1234 1235 1236 1237 1238 1239 1240 1241 1242 1243 1244 1245 1246 1247 1248 1249 1250 1251 1252 1253 1254 1255 1256 1257 1258 1259 1260 1261 1262 1263 1264 1265 1266 1267 1268 1269 1270 1271 1272 1273 1274 1275 1276 1277 1278 1279 1280 1281 1282 1283 1284 1285 1286 1287 1288 1289 1290 1291 1292 1293 1294 1295 1296 1297 1298 1299 1300 1301 1302 1303 1304 1305 1306 1307 1308 1309 1310 1311 1312 1313 1314 1315 1316 1317 1318 1319 1320 1321 1322 1323 1324 1325 1326 1327 1328 1329 1330 1331 1332 1333 1334 1335 1336 1337 1338 1339 1340 1341 1342 1343 1344 1345 1346 1347 1348 1349 1350 1351 1352 1353 1354 1355 1356 1357 1358 1359 1360 1361 1362 1363 1364 1365 1366 1367 1368 1369 1370 1371 1372 1373 1374 1375 1376 1377 1378 1379 1380 1381 1382 1383 1384 1385 1386 1387 1388 1389 1390 1391 1392 1393 1394 1395 1396 1397 1398 1399 1400 1401 1402 1403 1404 1405 1406 1407 1408 1409 1410 1411 1412 1413 1414 1415 1416 1417 1418 1419 1420 1421 1422 1423 1424 1425 1426 1427 1428 1429 1430 1431 1432 1433 1434 1435 1436 1437 1438 1439 1440 1441 1442 1443 1444 1445 1446 1447 1448 1449 1450 1451 1452 1453 1454 1455 1456 1457 1458 1459 1460 1461 1462 1463 1464 1465 1466 1467 1468 1469 1470 1471 1472 1473 1474 1475 1476 1477 1478 1479 1480 1481 1482 1483 1484 1485 1486 1487 1488 1489 1490 1491 1492 1493 1494 1495 1496 1497 1498 1499 1500 1501 1502 1503 1504 1505 1506 1507 1508 1509 1510 1511 1512 1513 1514 1515 1516 1517 1518 1519 1520 1521 1522 1523 1524 1525 1526 1527 1528 1529 1530 1531 1532 1533 1534 1535 1536 1537 1538 1539 1540 1541 1542 1543 1544 1545 1546 1547 1548 1549 1550 1551 1552 1553 1554 1555 1556 1557 1558 1559 1560 1561 1562 1563 1564 1565 1566 1567 1568 1569 1570 1571 1572 1573 1574 1575 1576 1577 1578 1579 1580 1581 1582 1583 1584 1585 1586 1587 1588 1589 1590 1591 1592 1593 1594 1595 1596 1597 1598 1599 1600 1601 1602 1603 1604 1605 1606 1607 1608 1609 1610 1611 1612 1613 1614 1615 1616 1617 1618 1619 1620 1621 1622 1623 1624 1625 1626 1627 1628 1629 1630 1631 1632 1633 1634 1635 1636 1637 1638 1639 1640 1641 1642 1643 1644 1645 1646 1647 1648 1649 1650 1651 1652 1653 1654 1655 1656 1657 1658 1659 1660 1661 1662 1663 1664 1665 1666 1667 1668 1669 1670 1671 1672 1673 1674 1675 1676 1677 1678 1679 1680 1681 1682 1683 1684 1685 1686 1687 1688 1689 1690 1691 1692 1693 1694 1695 1696 1697 1698 1699 1700 1701 1702 1703 1704 1705 1706 1707 1708 1709 1710 1711 1712 1713 1714 1715 1716 1717 1718 1719 1720 1721 1722 1723 1724 1725 1726 1727 1728 1729 1730 1731 1732 1733 1734 1735 1736 1737 1738 1739 1740 1741 1742 1743 1744 1745 1746 1747 1748 1749 1750 1751 1752 1753 1754 1755 1756 1757 1758 1759 1760 1761 1762 1763 1764 1765 1766 1767 1768 1769 1770 1771 1772 1773 1774 1775 1776 1777 1778 1779 1780 1781 1782 1783 1784 1785 1786 1787 1788 1789 1790 1791 1792 1793 1794 1795 1796 1797 1798 1799 1800 1801 1802 1803 1804 1805 1806 1807 1808 1809 1810 1811 1812 1813 1814 1815 1816 1817 1818 1819 1820 1821 1822 1823 1824 1825 1826 1827 1828 1829 1830 1831 1832 1833 1834 1835 1836 1837 1838 1839 1840 1841 1842 1843 1844 1845 1846 1847 1848 1849 1850 1851 1852 1853 1854 1855 1856 1857 1858 1859 1860 1861 1862 1863 1864 1865 1866 1867 1868 1869 1870 1871 1872 1873 1874 1875 1876 1877 1878 1879 1880 1881 1882 1883 1884 1885 1886 1887 1888 1889 1890 1891 1892 1893 1894 1895 1896 1897 1898 1899 1900 1901 1902 1903 1904 1905 1906 1907 1908 1909 1910 1911 1912 1913 1914 1915 1916 1917 1918 1919 1920 1921 1922 1923 1924 1925 1926 1927 1928 1929 1930 1931 1932 1933 1934 1935 1936 1937 1938 1939 1940 1941 1942 1943 1944 1945 1946 1947 1948 1949 1950 1951 1952 1953 1954 1955 1956 1957 1958 1959 1960 1961 1962 1963 1964 1965 1966 1967 1968 1969 1970 1971 1972 1973 1974 1975 1976 1977 1978 1979 1980 1981 1982 1983 1984 1985 1986 1987 1988 1989 1990 1991 1992 1993 1994 1995 1996 1997 1998 1999 2000 2001 2002 2003 2004 2005 2006 2007 2008 2009 2010 2011 2012 2013 2014 2015 2016 2017 2018 2019 2020 2021 2022 2023 2024 2025 2026 2027 2028 2029 2030 2031 2032 2033 2034 2035 2036 2037 2038 2039 2040 2041 2042 2043 2044 2045 2046 2047 2048 2049 2050 2051 2052 2053 2054 2055 2056 2057 2058 2059 2060 2061 2062 2063 2064 2065 2066 2067 2068 2069 2070 2071 2072 2073 2074 2075 2076 2077 2078 2079 2080 2081 2082 2083 2084 2085 2086 2087 2088 2089 2090 2091 2092 2093 2094 2095 2096 2097 2098 2099 2100 2101 2102 2103 2104 2105 2106 2107 2108 2109 2110 2111 2112 2113 2114 2115 2116 2117 2118 2119 2120 2121 2122 2123 2124 2125 2126 2127 2128 2129 2130 2131 2132 2133 2134 2135 2136 2137 2138 2139 2140 2141 2142 2143 2144 2145 2146 2147 2148 2149 2150 2151 2152 2153 2154 2155 2156 2157 2158 2159 2160 2161 2162 2163 2164 2165 2166 2167 2168 2169 2170 2171 2172 2173 2174 2175 2176 2177 2178 2179 2180 2181 2182 2183 2184 2185 2186 2187 2188 2189 2190 2191 2192 2193 2194 2195 2196 2197 2198 2199 2200 2201 2202 2203 2204 2205 2206 2207 2208 2209 2210 2211 2212 2213 2214 2215 2216 2217 2218 2219 2220 2221 2222 2223 2224 2225 2226 2227 2228 2229 2230 2231 2232 2233 2234 2235 2236 2237 2238 2239 2240 2241 2242 2243 2244 2245 2246 2247 2248 2249 2250 2251 2252 2253 2254 2255 2256 2257 2258 2259 2260 2261 2262 2263 2264 2265 2266 2267 2268 2269 2270 2271 2272 2273 2274 2275 2276 2277 2278 2279 2280 2281 2282 2283 2284 2285 2286 2287 2288 2289 2290 2291 2292 2293 2294 2295 2296 2297 2298 2299 2300 2301 2302 2303 2304 2305 2306 2307 2308 2309 2310 2311 2312 2313 2314 2315 2316 2317 2318 2319 2320 2321 2322 2323 2324 2325 2326 2327 2328 2329 2330 2331 2332 2333 2334 2335 2336 2337 2338 2339 2340 2341 2342 2343 2344 2345 2346 2347 2348 2349 2350 2351 2352 2353 2354 2355 2356 2357 2358 2359 2360 2361 2362 2363 2364 2365 2366 2367 2368 2369 2370 2371 2372 2373 2374 2375 2376 2377 2378 2379 2380 2381 2382 2383 2384 2385 2386 2387 2388 2389 2390 2391 2392 2393 2394 2395 2396 2397 2398 2399 2400 2401 2402 2403 2404 2405 2406 2407 2408 2409 2410 2411 2412 2413 2414 2415 2416 2417 2418 2419 2420 2421 2422 2423 2424 2425 2426 2427 2428 2429 2430 2431 2432 2433 2434 2435 2436 2437 2438 2439 2440 2441 2442 2443 2444 2445 2446 2447 2448 2449 2450 2451 2452 2453 2454 2455 2456 2457 2458 2459 2460 2461 2462 2463 2464 2465 2466 2467 2468 2469 2470 2471 2472 2473 2474 2475 2476 2477 2478 2479 2480 2481 2482 2483 2484 2485 2486 2487 2488 2489 2490 2491 2492 2493 2494 2495 2496 2497 2498 2499 2500 2501 2502 2503 2504 2505 2506 2507 2508 2509 2510 2511 2512 2513 2514 2515 2516 2517 2518 2519 2520 2521 2522 2523 2524 2525 2526 2527 2528 2529 2530 2531 2532 2533 2534 2535 2536 2537 2538 2539 2540 2541 2542 2543 2544 2545 2546 2547 2548 2549 2550 2551 2552 2553 2554 2555 2556 2557 2558 2559 2560 2561 2562 2563 2564 2565 2566 2567 2568 2569 2570 2571 2572 2573 2574 2575 2576 2577 2578 2579 2580 2581 2582 2583 2584 2585 2586 2587 2588 2589 2590 2591 2592 2593 2594 2595 2596 2597 2598 2599 2600 2601 2602 2603 2604 2605 2606 2607 2608 2609 2610 2611 2612 2613 2614 2615 2616 2617 2618 2619 2620 2621 2622 2623 2624 2625 2626 2627 2628 2629 2630 2631 2632 2633 2634 2635 2636 2637 2638 2639 2640 2641 2642 2643 2644 2645 2646 2647 2648 2649 2650 2651 2652 2653 2654 2655 2656 2657 2658 2659 2660 2661 2662 2663 2664 2665 2666 2667 2668 2669 2670 2671 2672 2673 2674 2675 2676 2677 2678 2679 2680 2681 2682 2683 2684 2685 2686 2687 2688 2689 2690 2691 2692 2693 2694 2695 2696 2697 2698 2699 2700 2701 2702 2703 2704 2705 2706 2707 2708 2709 2710 2711 2712 2713 2714 2715 2716 2717 2718 2719 2720 2721 2722 2723 2724 2725 2726 2727 2728 2729 2730 2731 2732 2733 2734 2735 2736 2737 2738 2739 2740 2741 2742 2743 2744 2745 2746 2747 2748 2749 2750 2751 2752 2753 2754 2755 2756 2757 2758 2759 2760 2761 2762 2763 2764 2765 2766 2767 2768 2769 2770 2771 2772 2773 2774 2775 2776 2777 2778 2779 2780 2781 2782 2783 2784 2785 2786 2787 2788 2789 2790 2791 2792 2793 2794 2795 2796 2797 2798 2799 2800 2801 2802 2803 2804 2805 2806 2807 2808 2809 2810 2811 2812 2813 2814 2815 2816 2817 2818 2819 2820 2821 2822 2823 2824 2825 2826 2827 2828 2829 2830 2831 2832 2833 2834 2835 2836 2837 2838 2839 2840 2841 2842 2843 2844 2845 2846 2847 2848 2849 2850 2851 2852 2853 2854 2855 2856 2857 2858 2859 2860 2861 2862 2863 2864 2865 2866 2867 2868 2869 2870 2871 2872 2873 2874 2875 2876 2877 2878 2879 2880 2881 2882 2883 2884 2885 2886 2887 2888 2889 2890 2891 2892 2893 2894 2895 2896 2897 2898 2899 2900 2901 2902 2903 2904 2905 2906 2907 2908 2909 2910 2911 2912 2913 2914 2915 2916 2917 2918 2919 2920 2921 2922 2923 2924 2925 2926 2927 2928 2929 2930 2931 2932 2933 2934 2935 2936 2937 2938 2939 2940 2941 2942 2943 2944 2945
|
#
# bioperl module for Bio::SeqFeature::Tools::Unflattener
#
# Please direct questions and support issues to <bioperl-l@bioperl.org>
#
# Cared for by Chris Mungall <cjm@fruitfly.org>
#
# Copyright Chris Mungall
#
# You may distribute this module under the same terms as perl itself
# POD documentation - main docs before the code
=head1 NAME
Bio::SeqFeature::Tools::Unflattener - turns flat list of genbank-sourced features into a nested SeqFeatureI hierarchy
=head1 SYNOPSIS
# standard / generic use - unflatten a genbank record
use Bio::SeqIO;
use Bio::SeqFeature::Tools::Unflattener;
# generate an Unflattener object
$unflattener = Bio::SeqFeature::Tools::Unflattener->new;
# first fetch a genbank SeqI object
$seqio =
Bio::SeqIO->new(-file=>'AE003644.gbk',
-format=>'GenBank');
my $out =
Bio::SeqIO->new(-format=>'asciitree');
while ($seq = $seqio->next_seq()) {
# get top level unflattended SeqFeatureI objects
$unflattener->unflatten_seq(-seq=>$seq,
-use_magic=>1);
$out->write_seq($seq);
@top_sfs = $seq->get_SeqFeatures;
foreach my $sf (@top_sfs) {
# do something with top-level features (eg genes)
}
}
=head1 DESCRIPTION
Most GenBank entries for annotated genomic DNA contain a B<flat> list
of features. These features can be parsed into an equivalent flat list
of L<Bio::SeqFeatureI> objects using the standard L<Bio::SeqIO>
classes. However, it is often desirable to B<unflatten> this list into
something resembling actual B<gene models>, in which genes, mRNAs and CDSs
are B<nested> according to the nature of the gene model.
The BioPerl object model allows us to store these kind of associations
between SeqFeatures in B<containment hierarchies> -- any SeqFeatureI
object can contain nested SeqFeatureI objects. The
Bio::SeqFeature::Tools::Unflattener object facilitates construction of
these hierarchies from the underlying GenBank flat-feature-list
representation.
For example, if you were to look at a typical GenBank DNA entry, say,
B<AE003644>, you would see a flat list of features:
source
gene CG4491
mRNA CG4491-RA
CDS CG4491-PA
gene tRNA-Pro
tRNA tRNA-Pro
gene CG32954
mRNA CG32954-RA
mRNA CG32954-RC
mRNA CG32954-RB
CDS CG32954-PA
CDS CG32954-PB
CDS CG32954-PC
These features have sequence locations, but it is not immediately
clear how to write code such that each mRNA is linked to the
appropriate CDS (other than relying on IDs which is very bad)
We would like to convert the above list into the B<containment
hierarchy>, shown below:
source
gene
mRNA CG4491-RA
CDS CG4491-PA
exon
exon
gene
tRNA tRNA-Pro
exon
gene
mRNA CG32954-RA
CDS CG32954-PA
exon
exon
mRNA CG32954-RC
CDS CG32954-PC
exon
exon
mRNA CG32954-RB
CDS CG32954-PB
exon
exon
Where each feature is nested underneath its container. Note that exons
have been automatically inferred (even for tRNA genes).
We do this using a call on a L<Bio::SeqFeature::Tools::Unflattener>
object
@sfs = $unflattener->unflatten_seq(-seq=>$seq);
This would return a list of the B<top level> (i.e. container)
SeqFeatureI objects - in this case, genes. Other top level features
are possible; for instance, the B<source> feature which is always
present, and other features such as B<variation> or B<misc_feature>
types.
The containment hierarchy can be accessed using the get_SeqFeature()
call on any feature object - see L<Bio::SeqFeature::FeatureHolderI>.
The following code will traverse the containment hierarchy for a
feature:
sub traverse {
$sf = shift; # $sf isa Bio::SeqfeatureI
# ...do something with $sf!
# depth first traversal of containment tree
@contained_sfs = $sf->get_SeqFeatures;
traverse($_) foreach @contained_sfs;
}
Once you have built the hierarchy, you can do neat stuff like turn the
features into 'rich' feature objects (eg
L<Bio::SeqFeature::Gene::GeneStructure>) or convert to a suitable
format such as GFF3 or chadoxml (after mapping to the Sequence
Ontology); this step is not described here.
=head1 USING MAGIC
Due to the quixotic nature of how features are stored in
GenBank/EMBL/DDBJ, there is no guarantee that the default behaviour of
this module will produce perfect results. Sometimes it is hard or
impossible to build a correct containment hierarchy if the information
provided is simply too lossy, as is often the case. If you care deeply
about your data, you should always manually inspect the resulting
containment hierarchy; you may have to customise the algorithm for
building the hierarchy, or even manually tweak the resulting
hierarchy. This is explained in more detail further on in the document.
However, if you are satisfied with the default behaviour, then you do
not need to read any further. Just make sure you set the parameter
B<use_magic> - this will invoke incantations which will magically
produce good results no matter what the idiosyncracies of the
particular GenBank record in question.
For example
$unflattener->unflatten_seq(-seq=>$seq,
-use_magic=>1);
The success of this depends on the phase of the moon at the time the
entry was submitted to GenBank. Note that the magical recipe is being
constantly improved, so the results of invoking magic may vary
depending on the bioperl release.
If you are skeptical of magic, or you wish to exact fine grained
control over how the entry is unflattened, or you simply wish to
understand more about how this crazy stuff works, then read on!
=head1 PROBLEMATIC DATA AND INCONSISTENCIES
Occasionally the Unflattener will have problems with certain
records. For example, the record may contain inconsistent data - maybe
there is an B<exon> entry that has no corresponding B<mRNA> location.
The default behaviour is to throw an exception reporting the problem,
if the problem is relatively serious - for example, inconsistent data.
You can exert more fine grained control over this - perhaps you want
the Unflattener to do the best it can, and report any problems. This
can be done - refer to the methods.
error_threshold()
get_problems()
report_problems()
ignore_problems()
=head1 ALGORITHM
This is the default algorithm; you should be able to override any part
of it to customise.
The core of the algorithm is in two parts
=over
=item Partitioning the flat feature list into groups
=item Resolving the feature containment hierarchy for each group
=back
There are other optional steps after the completion of these two
steps, such as B<inferring exons>; we now describe in more detail what
is going on.
=head2 Partitioning into groups
First of all the flat feature list is partitioned into B<group>s.
The default way of doing this is to use the B<gene> attribute; if we
look at two features from GenBank accession AE003644.3:
gene 20111..23268
/gene="noc"
/locus_tag="CG4491"
/note="last curated on Thu Dec 13 16:51:32 PST 2001"
/map="35B2-35B2"
/db_xref="FLYBASE:FBgn0005771"
mRNA join(20111..20584,20887..23268)
/gene="noc"
/locus_tag="CG4491"
/product="CG4491-RA"
/db_xref="FLYBASE:FBgn0005771"
Both these features share the same /gene tag which is "noc", so they
correspond to the same gene model (the CDS feature is not shown, but
this also has a tag-value /gene="noc").
Not all groups need to correspond to gene models, but this is the most
common use case; later on we shall describe how to customise the
grouping.
Sometimes other tags have to be used; for instance, if you look at the
entire record for AE003644.3 you will see you actually need the use the
/locus_tag attribute. This attribute is actually B<not present> in
most records!
You can override this:
$collection->unflatten_seq(-seq=>$seq, -group_tag=>'locus_tag');
Alternatively, if you B<-use_magic>, the object will try and make a
guess as to what the correct group_tag should be.
At the end of this step, we should have a list of groups - there is no
structure within a group; the group just serves to partition the flat
features. For the example data above, we would have the following groups.
[ source ]
[ gene mRNA CDS ]
[ gene mRNA CDS ]
[ gene mRNA CDS ]
[ gene mRNA mRNA mRNA CDS CDS CDS ]
=head3 Multicopy Genes
Multicopy genes are usually rRNAs or tRNAs that are duplicated across
the genome. Because they are functionally equivalent, and usually have
the same sequence, they usually have the same group_tag (ie gene
symbol); they often have a /note tag giving copy number. This means
they will end up in the same group. This is undesirable, because they
are spatially disconnected.
There is another step, which involves splitting spatially disconnected
groups into distinct groups
this would turn this
[gene-rrn3 rRNA-rrn3 gene-rrn3 rRNA-rrn3]
into this
[gene-rrn3 rRNA-rrn3] [gene-rrn3 rRNA-rrn3]
based on the coordinates
=head3 What next?
The next step is to add some structure to each group, by making
B<containment hierarchies>, trees that represent how the features
interrelate
=head2 Resolving the containment hierarchy
After the grouping is done, we end up with a list of groups which
probably contain features of type 'gene', 'mRNA', 'CDS' and so on.
Singleton groups (eg the 'source' feature) are ignored at this stage.
Each group is itself flat; we need to add an extra level of
organisation. Usually this is because different spliceforms
(represented by the 'mRNA' feature) can give rise to different
protein products (indicated by the 'CDS' feature). We want to correctly
associate mRNAs to CDSs.
We want to go from a group like this:
[ gene mRNA mRNA mRNA CDS CDS CDS ]
to a containment hierarchy like this:
gene
mRNA
CDS
mRNA
CDS
mRNA
CDS
In which each CDS is nested underneath the correct corresponding mRNA.
For entries that contain no alternate splicing, this is simple; we
know that the group
[ gene mRNA CDS ]
Must resolve to the tree
gene
mRNA
CDS
How can we do this in entries with alternate splicing? The bad
news is that there is no guaranteed way of doing this correctly for
any GenBank entry. Occasionally the submission will have been done in
such a way as to reconstruct the containment hierarchy. However, this
is not consistent across databank entries, so no generic solution can
be provided by this object. This module does provide the framework
within which you can customise a solution for the particular dataset
you are interested in - see later.
The good news is that there is an inference we can do that should
produce pretty good results the vast majority of the time. It uses
splice coordinate data - this is the default behaviour of this module,
and is described in detail below.
=head2 Using splice site coordinates to infer containment
If an mRNA is to be the container for a CDS, then the splice site
coordinates (or intron coordinates, depending on how you look at it)
of the CDS must fit inside the splice site coordinates of the mRNA.
Ambiguities can still arise, but the results produced should still be
reasonable and consistent at the sequence level. Look at this fake
example:
mRNA XXX---XX--XXXXXX--XXXX join(1..3,7..8,11..16,19..23)
mRNA XXX-------XXXXXX--XXXX join(1..3,11..16,19..23)
CDS XXXX--XX join(13..16,19..20)
CDS XXXX--XX join(13..16,19..20)
[obviously the positions have been scaled down]
We cannot unambiguously match mRNA with CDS based on splice sites,
since both CDS share the splice site locations 16^17 and
18^19. However, the consequences of making a wrong match are probably
not very severe. Any annotation data attached to the first CDS is
probably identical to the seconds CDS, other than identifiers.
The default behaviour of this module is to make an arbitrary call
where it is ambiguous (the mapping will always be bijective; i.e. one
mRNA -E<gt> one CDS).
[TODO: NOTE: not tested on EMBL data, which may not be bijective; ie two
mRNAs can share the same CDS??]
This completes the building of the containment hierarchy; other
optional step follow
=head1 POST-GROUPING STEPS
=head2 Inferring exons from mRNAs
This step always occurs if B<-use_magic> is invoked.
In a typical GenBank entry, the exons are B<implicit>. That is they
can be inferred from the mRNA location.
For example:
mRNA join(20111..20584,20887..23268)
This tells us that this particular transcript has two exons. In
bioperl, the mRNA feature will have a 'split location'.
If we call
$unflattener->feature_from_splitloc(-seq=>$seq);
This will generate the necessary exon features, and nest them under
the appropriate mRNAs. Note that the mRNAs will no longer have split
locations - they will have simple locations spanning the extent of the
exons. This is intentional, to avoid redundancy.
Occasionally a GenBank entry will have both implicit exons (from the
mRNA location) B<and> explicit exon features.
In this case, exons will still be transferred. Tag-value data from the
explicit exon will be transfered to the implicit exon. If exons are
shared between mRNAs these will be represented by different
objects. Any inconsistencies between implicit and explicit will be
reported.
=head3 tRNAs and other noncoding RNAs
exons will also be generated from these features
=head2 Inferring mRNAs from CDS
Some GenBank entries represent gene models using features of type
gene, mRNA and CDS; some entries just use gene and CDS.
If we only have gene and CDS, then the containment hierarchies will
look like this:
gene
CDS
If we want the containment hierarchies to be uniform, like this
gene
mRNA
CDS
Then we must create an mRNA feature. This will have identical
coordinates to the CDS. The assumption is that there is either no
untranslated region, or it is unknown.
To do this, we can call
$unflattener->infer_mRNA_from_CDS(-seq=>$seq);
This is taken care of automatically, if B<-use_magic> is invoked.
=head1 ADVANCED
=head2 Customising the grouping of features
The default behaviour is suited mostly to building models of protein
coding genes and noncoding genes from genbank genomic DNA submissions.
You can change the tag used to partition the feature by passing in a
different group_tag argument - see the unflatten_seq() method
Other behaviour may be desirable. For example, even though SNPs
(features of type 'variation' in GenBank) are not actually part of the
gene model, it may be desirable to group SNPs that overlap or are
nearby gene models.
It should certainly be possible to extend this module to do
this. However, I have yet to code this part!!! If anyone would find
this useful let me know.
In the meantime, you could write your own grouping subroutine, and
feed the results into unflatten_groups() [see the method documentation
below]
=head2 Customising the resolution of the containment hierarchy
Once the flat list of features has been partitioned into groups, the
method unflatten_group() is called on each group to build a tree.
The algorithm for doing this is described above; ambiguities are
resolved by using splice coordinates. As discussed, this can be
ambiguous.
Some submissions may contain information in tags/attributes that hint
as to the mapping that needs to be made between the features.
For example, with the Drosophila Melanogaster release 3 submission, we
see that CDS features in alternately spliced mRNAs have a form like
this:
CDS join(145588..145686,145752..146156,146227..146493)
/locus_tag="CG32954"
/note="CG32954 gene product from transcript CG32954-RA"
^^^^^^^^^^^^^^^^^^^^^^^^^^^
/codon_start=1
/product="CG32954-PA"
/protein_id="AAF53403.1"
/db_xref="GI:7298167"
/db_xref="FLYBASE:FBgn0052954"
/translation="MSFTLTNKNVIFVAGLGGIGLDTSKELLKRDLKNLVILDRIENP..."
Here the /note tag provides the clue we need to link CDS to mRNA
(highlighted with ^^^^). We just need to find the mRNA with the tag
/product="CG32954-RA"
I have no idea how consistent this practice is across submissions; it
is consistent for the fruitfly genome submission.
We can customise the behaviour of unflatten_group() by providing our
own resolver method. This obviously requires a bit of extra
programming, but there is no way to get around this.
Here is an example of how to pass in your own resolver; this example
basically checks the parent (container) /product tag to see if it
matches the required string in the child (contained) /note tag.
$unflattener->unflatten_seq(-seq=>$seq,
-group_tag=>'locus_tag',
-resolver_method=>sub {
my $self = shift;
my ($sf, @candidate_container_sfs) = @_;
if ($sf->has_tag('note')) {
my @notes = $sf->get_tag_values('note');
my @trnames = map {/from transcript\s+(.*)/;
$1} @notes;
@trnames = grep {$_} @trnames;
my $trname;
if (@trnames == 0) {
$self->throw("UNRESOLVABLE");
}
elsif (@trnames == 1) {
$trname = $trnames[0];
}
else {
$self->throw("AMBIGUOUS: @trnames");
}
my @container_sfs =
grep {
my ($product) =
$_->has_tag('product') ?
$_->get_tag_values('product') :
('');
$product eq $trname;
} @candidate_container_sfs;
if (@container_sfs == 0) {
$self->throw("UNRESOLVABLE");
}
elsif (@container_sfs == 1) {
# we got it!
return $container_sfs[0];
}
else {
$self->throw("AMBIGUOUS");
}
}
});
the resolver method is only called when there is more than one spliceform.
=head2 Parsing mRNA records
Some of the entries in sequence databanks are for mRNA sequences as
well as genomic DNA. We may want to build models from these too.
NOT YET DONE - IN PROGRESS!!!
Open question - what would these look like?
Ideally we would like a way of combining a mRNA record with the
corresponding SeFeature entry from the appropriate genomic DNA
record. This could be problemmatic in some cases - for example, the
mRNA sequences may not match 100% (due to differences in strain,
assembly problems, sequencing problems, etc). What then...?
=head1 SEE ALSO
Feature table description
http://www.ebi.ac.uk/embl/Documentation/FT_definitions/feature_table.html
=head1 FEEDBACK
=head2 Mailing Lists
User feedback is an integral part of the evolution of this and other
Bioperl modules. Send your comments and suggestions preferably to the
Bioperl mailing lists Your participation is much appreciated.
bioperl-l@bioperl.org - General discussion
http://bioperl.org/wiki/Mailing_lists - About the mailing lists
=head2 Support
Please direct usage questions or support issues to the mailing list:
I<bioperl-l@bioperl.org>
rather than to the module maintainer directly. Many experienced and
reponsive experts will be able look at the problem and quickly
address it. Please include a thorough description of the problem
with code and data examples if at all possible.
=head2 Reporting Bugs
report bugs to the Bioperl bug tracking system to help us keep track
the bugs and their resolution. Bug reports can be submitted via the
web:
https://github.com/bioperl/bioperl-live/issues
=head1 AUTHOR - Chris Mungall
Email: cjm@fruitfly.org
=head1 APPENDIX
The rest of the documentation details each of the object
methods. Internal methods are usually preceded with a _
=cut
# Let the code begin...
package Bio::SeqFeature::Tools::Unflattener;
use strict;
# Object preamble - inherits from Bio::Root::Root
use Bio::Location::Simple;
use Bio::SeqFeature::Generic;
use Bio::Range;
use base qw(Bio::Root::Root);
=head2 new
Title : new
Usage : $unflattener = Bio::SeqFeature::Tools::Unflattener->new();
$unflattener->unflatten_seq(-seq=>$seq);
Function: constructor
Example :
Returns : a new Bio::SeqFeature::Tools::Unflattener
Args : see below
Arguments
-seq : A L<Bio::SeqI> object (optional)
the sequence to unflatten; this can also be passed in
when we call unflatten_seq()
-group_tag : a string representing the /tag used to partition flat features
(see discussion above)
=cut
sub new {
my($class,@args) = @_;
my $self = $class->SUPER::new(@args);
my($seq, $group_tag, $trust_grouptag) =
$self->_rearrange([qw(SEQ
GROUP_TAG
TRUST_GROUPTAG
)],
@args);
$seq && $self->seq($seq);
$group_tag && $self->group_tag($group_tag);
# $self->{'trust_grouptag'}= $trust_grouptag if($trust_grouptag); #dgg suggestion
return $self; # success - we hope!
}
sub DESTROY {
my $self = shift;
return if $self->{_reported_problems};
return if $self->{_ignore_problems};
my @probs = $self->get_problems;
if (!$self->{_problems_reported} &&
scalar(@probs)) {
$self->warn(
"WARNING: There are UNREPORTED PROBLEMS.\n".
"You may wish to use the method report_problems(), \n",
"or ignore_problems() on the Unflattener object\n");
}
return;
}
=head2 seq
Title : seq
Usage : $unflattener->seq($newval)
Function:
Example :
Returns : value of seq (a Bio::SeqI)
Args : on set, new value (a Bio::SeqI, optional)
The Bio::SeqI object should hold a flat list of Bio::SeqFeatureI
objects; this is the list that will be unflattened.
The sequence object can also be set when we call unflatten_seq()
=cut
sub seq{
my $self = shift;
return $self->{'seq'} = shift if @_;
return $self->{'seq'};
}
=head2 group_tag
Title : group_tag
Usage : $unflattener->group_tag($newval)
Function:
Example :
Returns : value of group_tag (a scalar)
Args : on set, new value (a scalar or undef, optional)
This is the tag that will be used to collect elements from the flat
feature list into groups; for instance, if we look at two typical
GenBank features:
gene 20111..23268
/gene="noc"
/locus_tag="CG4491"
/note="last curated on Thu Dec 13 16:51:32 PST 2001"
/map="35B2-35B2"
/db_xref="FLYBASE:FBgn0005771"
mRNA join(20111..20584,20887..23268)
/gene="noc"
/locus_tag="CG4491"
/product="CG4491-RA"
/db_xref="FLYBASE:FBgn0005771"
We can see that these comprise the same gene model because they share
the same /gene attribute; we want to collect these together in groups.
Setting group_tag is optional. The default is to use 'gene'. In the
example above, we could also use /locus_tag
=cut
sub group_tag{
my $self = shift;
return $self->{'group_tag'} = shift if @_;
return $self->{'group_tag'};
}
=head2 partonomy
Title : partonomy
Usage : $unflattener->partonomy({mRNA=>'gene', CDS=>'mRNA')
Function:
Example :
Returns : value of partonomy (a scalar)
Args : on set, new value (a scalar or undef, optional)
A hash representing the containment structure that the seq_feature
nesting should conform to; each key represents the contained (child)
type; each value represents the container (parent) type.
=cut
sub partonomy{
my $self = shift;
return $self->{'partonomy'} = shift if @_;
if (!$self->{'partonomy'}) {
$self->{'partonomy'} = $self->_default_partonomy;
}
return $self->{'partonomy'};
}
sub _default_partonomy{
return {
mRNA => 'gene',
tRNA => 'gene',
rRNA => 'gene',
scRNA => 'gene',
snRNA => 'gene',
snoRNA => 'gene',
misc_RNA => 'gene',
CDS => 'mRNA',
exon => 'mRNA',
intron => 'mRNA',
pseudoexon => 'pseudogene',
pseudointron => 'pseudogene',
pseudotranscript => 'pseudogene',
};
}
=head2 structure_type
Title : structure_type
Usage : $unflattener->structure_type($newval)
Function:
Example :
Returns : value of structure_type (a scalar)
Args : on set, new value (an int or undef, optional)
GenBank entries conform to different flavours, or B<structure
types>. Some have mRNAs, some do not.
Right now there are only two base structure types defined. If you set
the structure type, then appropriate unflattening action will be
taken. The presence or absence of explicit exons does not affect the
structure type.
If you invoke B<-use_magic> then this will be set automatically, based
on the content of the record.
=over
=item Type 0 (DEFAULT)
typically contains
source
gene
mRNA
CDS
with this structure type, we want the seq_features to be nested like this
gene
mRNA
CDS
exon
exons and introns are implicit from the mRNA 'join' location
to get exons from the mRNAs, you will need this call (see below)
$unflattener->feature_from_splitloc(-seq=>$seq);
=item Type 1
typically contains
source
gene
CDS
exon [optional]
intron [optional]
there are no mRNA features
with this structure type, we want the seq_features to be nested like this
gene
CDS
exon
intron
exon and intron may or may not be present; they may be implicit from
the CDS 'join' location
=back
=cut
sub structure_type{
my $self = shift;
return $self->{'structure_type'} = shift if @_;
return $self->{'structure_type'};
}
=head2 get_problems
Title : get_problems
Usage : @probs = get_problems()
Function: Get the list of problem(s) for this object.
Example :
Returns : An array of [severity, description] pairs
Args :
In the course of unflattening a record, problems may occur. Some of
these problems are non-fatal, and can be ignored.
Problems are represented as arrayrefs containing a pair [severity,
description]
severity is a number, the higher, the more severe the problem
the description is a text string
=cut
sub get_problems{
my $self = shift;
return @{$self->{'_problems'}} if exists($self->{'_problems'});
return ();
}
=head2 clear_problems
Title : clear_problems
Usage :
Function: resets the problem list to empty
Example :
Returns :
Args :
=cut
sub clear_problems{
my ($self,@args) = @_;
$self->{'_problems'} = [];
return;
}
# PRIVATE
# see get_problems
sub add_problem{
my $self = shift;
$self->{'_problems'} = [] unless exists($self->{'_problems'});
if ($self->verbose > 0) {
warn( "PROBLEM: $_\n") foreach @_;
}
push(@{$self->{'_problems'}}, @_);
}
# PRIVATE
# see get_problems
sub problem {
my $self = shift;
my ($severity, $desc, @sfs) = @_;
if (@sfs) {
foreach my $sf (@sfs) {
$desc .=
sprintf("\nSF [$sf]: ". $sf->location->to_FTstring . "; %s\n",
join('; ',
$sf->primary_tag,
map {
$sf->has_tag($_) ?
$sf->get_tag_values($_) : ()
} qw(locus_tag gene product label)));
}
}
my $thresh = $self->error_threshold;
if ($severity > $thresh) {
$self->{_problems_reported} = 1;
$self->throw("PROBLEM, SEVERITY==$severity\n$desc");
}
$self->add_problem([$severity, $desc]);
return;
}
=head2 report_problems
Title : report_problems
Usage : $unflattener->report_problems(\*STDERR);
Function:
Example :
Returns :
Args : FileHandle (defaults to STDERR)
=cut
sub report_problems{
my ($self, $fh) = @_;
if (!$fh) {
$fh = \*STDERR;
}
foreach my $problem ($self->get_problems) {
my ($sev, $desc) = @$problem;
printf $fh "PROBLEM, SEVERITY==$sev\n$desc\n";
}
$self->{_problems_reported} = 1;
return;
}
=head2 ignore_problems
Title : ignore_problems
Usage : $obj->ignore_problems();
Function:
Example :
Returns :
Args :
Unflattener is very particular about problems it finds along the
way. If you have set the error_threshold such that less severe
problems do not cause exceptions, Unflattener still expects you to
report_problems() at the end, so that the user of the module is aware
of any inconsistencies or problems with the data. In fact, a warning
will be produced if there are unreported problems. To silence, this
warning, call the ignore_problems() method before the Unflattener
object is destroyed.
=cut
sub ignore_problems{
my ($self) = @_;
$self->{_ignore_problems} = 1;
return;
}
=head2 error_threshold
Title : error_threshold
Usage : $obj->error_threshold($severity)
Function:
Example :
Returns : value of error_threshold (a scalar)
Args : on set, new value (an integer)
Sets the threshold above which errors cause this module to throw an
exception. The default is 0; all problems with a severity E<gt> 0 will
cause an exception.
If you raise the threshold to 1, then the unflattening process will be
more lax; problems of severity==1 are generally non-fatal, but may
indicate that the results should be inspected, for example, to make
sure there is no data loss.
=cut
sub error_threshold{
my $self = shift;
return $self->{'error_threshold'} = shift if @_;
return $self->{'error_threshold'} || 0;
}
# PRIVATE
#
# given a type (eg mRNA), will return the container type (eg gene)
sub get_container_type{
my ($self,$type) = @_;
my @roots = $self->_get_partonomy_roots;
if (grep {$_ eq $type} @roots) {
# it is a root - no parents/containers
return;
}
my $ch = $self->partonomy;
my $ctype = $ch->{$type};
if (!$ctype) {
# asterix acts as a wild card
$ctype = $ch->{'*'};
}
return $ctype;
}
# get root node of partonomy hierarchy (usually gene)
sub _get_partonomy_roots {
my $self = shift;
my $ch = $self->partonomy;
my @parents = values %$ch;
# find parents that do not have parents themselves
return grep {!$ch->{$_}} @parents;
}
=head2 unflatten_seq
Title : unflatten_seq
Usage : @sfs = $unflattener->unflatten_seq($seq);
Function: turns a flat list of features into a list of holder features
Example :
Returns : list of Bio::SeqFeatureI objects
Args : see below
partitions a list of features then arranges them in a nested tree; see
above for full explanation.
note - the Bio::SeqI object passed in will be modified
Arguments
-seq : a Bio::SeqI object; must contain Bio::SeqFeatureI objects
(this is optional if seq has already been set)
-use_magic: if TRUE (ie non-zero) then magic will be invoked;
see discussion above.
-resolver_method: a CODE reference
see the documentation above for an example of
a subroutine that can be used to resolve hierarchies
within groups.
this is optional - if nothing is supplied, a default
subroutine will be used (see below)
-group_tag: a string
[ see the group_tag() method ]
this overrides the default group_tag which is 'gene'
=cut
sub unflatten_seq{
my ($self,@args) = @_;
my($seq, $resolver_method, $group_tag, $partonomy,
$structure_type, $resolver_tag, $use_magic, $noinfer) =
$self->_rearrange([qw(SEQ
RESOLVER_METHOD
GROUP_TAG
PARTONOMY
STRUCTURE_TYPE
RESOLVER_TAG
USE_MAGIC
NOINFER
)],
@args);
# seq we want to unflatten
$seq = $seq || $self->seq;
if (!$self->seq) {
$self->seq($seq);
}
# prevent bad argument combinations
if ($partonomy &&
defined($structure_type)) {
$self->throw("You cannot set both -partonomy and -structure_type\n".
"(the former is implied by the latter)");
}
# remember the current value of partonomy, to reset later
my $old_partonomy = $self->partonomy;
$self->partonomy($partonomy) if defined $partonomy;
# remember old structure_type
my $old_structure_type = $self->structure_type;
$self->structure_type($structure_type) if defined $structure_type;
# if we are sourcing our data from genbank, all the
# features should be flat (eq no sub_SeqFeatures)
my @flat_seq_features = $seq->get_SeqFeatures;
my @all_seq_features = $seq->get_all_SeqFeatures;
# sanity checks
if (@all_seq_features > @flat_seq_features) {
$self->throw("It looks as if this sequence has already been unflattened");
}
if (@all_seq_features < @flat_seq_features) {
$self->throw("ASSERTION ERROR: something is seriously wrong with your features");
}
# tag for ungrouping; usually /gene or /locus_tag
# for example: /gene="foo"
$group_tag = $group_tag || $self->group_tag;
if ($use_magic) {
# use magic to guess the group tag
my @sfs_with_locus_tag =
grep {$_->has_tag("locus_tag")} @flat_seq_features;
my @sfs_with_gene_tag =
grep {$_->has_tag("gene")} @flat_seq_features;
my @sfs_with_product_tag =
grep {$_->has_tag("product")} @flat_seq_features;
# if ($group_tag && $self->{'trust_grouptag'}) { # dgg suggestion
#
# }
# elsif
if (@sfs_with_locus_tag) {
# dgg note: would like to -use_magic with -group_tag = 'gene' for ensembl genomes
# where ensembl gene FT have both /locus_tag and /gene, but mRNA, CDS have /gene only
if ($group_tag && $group_tag ne 'locus_tag') {
$self->throw("You have explicitly set group_tag to be '$group_tag'\n".
"However, I detect that some features use /locus_tag\n".
"I believe that this is the correct group_tag to use\n".
"You can resolve this by either NOT setting -group_tag\n".
"OR you can unset -use_magic to regain control");
}
# use /locus_tag instead of /gene tag for grouping
# see GenBank entry AE003677 (version 3) for an example
$group_tag = 'locus_tag';
if ($self->verbose > 0) {
warn "Set group tag to: $group_tag\n";
}
}
# on rare occasions, records will have no /gene or /locus_tag
# but it WILL have /product tags. These serve the same purpose
# for grouping. For an example, see AY763288 (also in t/data)
if (@sfs_with_locus_tag==0 &&
@sfs_with_gene_tag==0 &&
@sfs_with_product_tag>0 &&
!$group_tag) {
$group_tag = 'product';
if ($self->verbose > 0) {
warn "Set group tag to: $group_tag\n";
}
}
}
if (!$group_tag) {
$group_tag = 'gene';
}
# ------------------------------
# GROUP FEATURES using $group_tag
# collect features into unstructured groups
# ------------------------------
# -------------
# we want to generate a list of groups;
# each group is a list of SeqFeatures; this
# group probably (but not necessarily)
# corresponds to a gene model.
#
# this array will look something like this:
# ([$f1], [$f2, $f3, $f4], ...., [$f97, $f98, $f99])
#
# there are also 'singleton' groups, with one member.
# for instance, the 'source' feature is in a singleton group;
# the same with others such as 'misc_feature'
my @groups = ();
# -------------
# --------------------
# we hope that the genbank record allows us to group by some grouping
# tag.
# for instance, most of the time a gene model can be grouped using
# the gene tag - that is where you see
# /gene="foo"
# in a genbank record
# --------------------
# keep an index of groups by their
# grouping tag
my %group_by_tag = ();
# iterate through all features, putting them into groups
foreach my $sf (@flat_seq_features) {
if (!$sf->has_tag($group_tag)) {
# SINGLETON
# this is an ungroupable feature;
# add it to a group of its own
push(@groups, [$sf]);
}
else {
# NON-SINGLETON
my @group_tagvals = $sf->get_tag_values($group_tag);
if (@group_tagvals > 1) {
# sanity check:
# currently something can only belong to one group
$self->problem(2,
">1 value for /$group_tag: @group_tagvals\n".
"At this time this module is not equipped to handle this adequately", $sf);
}
# get value of group tag
my $gtv = shift @group_tagvals;
$gtv || $self->throw("Empty /$group_tag vals not allowed!");
# is this a new group?
my $group = $group_by_tag{$gtv};
if ($group) {
# this group has been encountered before - add current
# sf to the end of the group
push(@$group, $sf);
}
else {
# new group; add to index and create new group
$group = [$sf]; # currently one member; probably more to come
$group_by_tag{$gtv} = $group;
push(@groups, $group);
}
}
}
# as well as having the same group_tag, a group should be spatially
# connected. if not, then the group should be split into subgroups.
# this turns out to be necessary in the case of multicopy genes.
# the standard way to represent these is as spatially disconnected
# gene models (usually a 'gene' feature and some kind of RNA feature)
# with the same group tag; the code below will split these into
# seperate groups, one per copy.
@groups = map { $self->_split_group_if_disconnected($_) } @groups;
# remove any duplicates; most of the time the method below has
# no effect. there are some unusual genbank records for which
# duplicate removal is necessary. see the comments in the
# _remove_duplicates_from_group() method if you want to know
# the ugly details
foreach my $group (@groups) {
$self->_remove_duplicates_from_group($group);
}
# -
# PSEUDOGENES, PSEUDOEXONS AND PSEUDOINTRONS
# these are indicated with the /pseudo tag
# these are mapped to a different type; they should NOT
# be treated as normal genes
foreach my $sf (@all_seq_features) {
if ($sf->has_tag('pseudo')) {
my $type = $sf->primary_tag;
# SO type is typically the same as the normal
# type but preceeded by "pseudo"
if ($type eq 'misc_RNA' || $type eq 'mRNA') {
# dgg: see TypeMapper; both pseudo mRNA,misc_RNA should be pseudogenic_transcript
$sf->primary_tag("pseudotranscript");
}
else {
$sf->primary_tag("pseudo$type");
}
}
}
# now some of the post-processing that follows which applies to
# genes will NOT be applied to pseudogenes; this is deliberate
# for example, gene models are normalised to be gene-transcript-exon
# for pseudogenes we leave them as pseudogene-pseudoexon
# --- MAGIC ---
my $need_to_infer_exons = 0;
my $need_to_infer_mRNAs = 0;
my @removed_exons = ();
if ($use_magic) {
if (defined($structure_type)) {
$self->throw("Can't combine use_magic AND setting structure_type");
}
my $n_introns =
scalar(grep {$_->primary_tag eq 'exon'} @flat_seq_features);
my $n_exons =
scalar(grep {$_->primary_tag eq 'exon'} @flat_seq_features);
my $n_mrnas =
scalar(grep {$_->primary_tag eq 'mRNA'} @flat_seq_features);
my $n_mrnas_attached_to_gene =
scalar(grep {$_->primary_tag eq 'mRNA' &&
$_->has_tag($group_tag)} @flat_seq_features);
my $n_cdss =
scalar(grep {$_->primary_tag eq 'CDS'} @flat_seq_features);
my $n_rnas =
scalar(grep {$_->primary_tag =~ /RNA/} @flat_seq_features);
# Are there any CDS features in the record?
if ($n_cdss > 0) {
# YES
# - a pc gene model should contain at the least a CDS
# Are there any mRNA features in the record?
if ($n_mrnas == 0) {
# NO mRNAs:
# looks like structure_type == 1
$structure_type = 1;
$need_to_infer_mRNAs = 1;
}
elsif ($n_mrnas_attached_to_gene == 0) {
# $n_mrnas > 0
# $n_mrnas_attached_to_gene = 0
#
# The entries _do_ contain mRNA features,
# but none of them are part of a group/gene, i.e. they
# are 'floating'
# this is an annoying weird file that has some floating
# mRNA features;
# eg ftp.ncbi.nih.gov/genomes/Schizosaccharomyces_pombe/
if ($self->verbose) {
my @floating_mrnas =
grep {$_->primary_tag eq 'mRNA' &&
!$_->has_tag($group_tag)} @flat_seq_features;
printf STDERR "Unattached mRNAs:\n";
foreach my $mrna (@floating_mrnas) {
$self->_write_sf_detail($mrna);
}
printf STDERR "Don't know how to deal with these; filter at source?\n";
}
foreach (@flat_seq_features) {
if ($_->primary_tag eq 'mRNA') {
# what should we do??
# I think for pombe we just have to filter
# out bogus mRNAs prior to starting
}
}
# looks like structure_type == 2
$structure_type = 2;
$need_to_infer_mRNAs = 1;
}
else {
}
# we always infer exons in magic mode
$need_to_infer_exons = 1;
}
else {
# this doesn't seem to be any kind of protein coding gene model
if ( $n_rnas > 0 ) {
$need_to_infer_exons = 1;
}
}
$need_to_infer_exons = 0 if $noinfer; #NML
if ($need_to_infer_exons) {
# remove exons and introns from group -
# we will infer exons later, and we
# can always infer introns from exons
foreach my $group (@groups) {
@$group =
grep {
my $type = $_->primary_tag();
if ($type eq 'exon') {
# keep track of all removed exons,
# so we can do a sanity check later
push(@removed_exons, $_);
}
$type ne 'exon' && $type ne 'intron'
} @$group;
}
# get rid of any groups that have zero members
@groups = grep {scalar(@$_)} @groups;
}
}
# --- END OF MAGIC ---
# LOGICAL ASSERTION
if (grep {!scalar(@$_)} @groups) {
$self->throw("ASSERTION ERROR: empty group");
}
# LOGGING
if ($self->verbose > 0) {
printf STDERR "GROUPS:\n";
foreach my $group (@groups) {
$self->_write_group($group, $group_tag);
}
}
# -
# --------- FINISHED GROUPING -------------
# TYPE CONTAINMENT HIERARCHY (aka partonomy)
# set the containment hierarchy if desired
# see docs for structure_type() method
if ($structure_type) {
if ($structure_type == 1) {
$self->partonomy(
{CDS => 'gene',
exon => 'CDS',
intron => 'CDS',
}
);
}
else {
$self->throw("structure_type $structure_type is currently unknown");
}
}
# see if we have an obvious resolver_tag
if ($use_magic) {
foreach my $sf (@all_seq_features) {
if ($sf->has_tag('derived_from')) {
$resolver_tag = 'derived_from';
}
}
}
if ($use_magic) {
# point all feature types without a container type to the root type.
#
# for example, if we have an unanticipated feature_type, say
# 'aberration', this should by default point to the parent 'gene'
foreach my $group (@groups) {
my @sfs = @$group;
if (@sfs > 1) {
foreach my $sf (@sfs) {
my $type = $sf->primary_tag;
next if $type eq 'gene';
my $container_type = $self->get_container_type($type);
if (!$container_type) {
$self->partonomy->{$type} = 'gene';
}
}
}
}
}
# we have done the first part of the unflattening.
# we now have a list of groups; each group is a list of seqfeatures.
# the actual group itself is flat; we may want to unflatten this further;
# for instance, a gene model can contain multiple mRNAs and CDSs. We may want
# to link the correct mRNA to the correct CDS via the bioperl sub_SeqFeature tree.
#
# what we would end up with would be
# gene1
# mRNA-a
# CDS-a
# mRNA-b
# CDS-b
my @top_sfs = $self->unflatten_groups(-groups=>\@groups,
-resolver_method=>$resolver_method,
-resolver_tag=>$resolver_tag);
# restore settings
$self->partonomy($old_partonomy);
# restore settings
$self->structure_type($old_structure_type);
# modify the original Seq object - the top seqfeatures are now
# the top features from each group
$seq->remove_SeqFeatures;
$seq->add_SeqFeature($_) foreach @top_sfs;
# --------- FINISHED UNFLATTENING -------------
# lets see if there are any post-unflattening tasks we need to do
# INFERRING mRNAs
if ($need_to_infer_mRNAs) {
if ($self->verbose > 0) {
printf STDERR "** INFERRING mRNA from CDS\n";
}
$self->infer_mRNA_from_CDS(-seq=>$seq, -noinfer=>$noinfer);
}
# INFERRING exons
if ($need_to_infer_exons) {
# infer exons, one group/gene at a time
foreach my $sf (@top_sfs) {
my @sub_sfs = ($sf, $sf->get_all_SeqFeatures);
$self->feature_from_splitloc(-features=>\@sub_sfs);
}
# some exons are stated explicitly; ie there is an "exon" feature
# most exons are inferred; ie there is a "mRNA" feature with
# split locations
#
# if there were exons explicitly stated in the entry, we need to
# do two things:
#
# make sure these exons are consistent with the inferred exons
# (you never know)
#
# transfer annotation (tag-vals) from the explicit exon to the
# new inferred exon
if (@removed_exons) {
my @allfeats = $seq->get_all_SeqFeatures;
# find all the inferred exons that are children of mRNA
my @mrnas = grep {$_->primary_tag eq 'mRNA'} @allfeats;
my @exons =
grep {$_->primary_tag eq 'exon'}
map {$_->get_SeqFeatures} @mrnas;
my %exon_h = (); # index of exons by location;
# there CAN be >1 exon at a location; we can represent these redundantly
# (ie as a tree, not a graph)
push(@{$exon_h{$self->_locstr($_)}}, $_) foreach @exons;
my @problems = (); # list of problems;
# each problem is a
# [$severity, $description] pair
my $problem = '';
my ($n_exons, $n_removed_exons) =
(scalar(keys %exon_h), scalar(@removed_exons));
foreach my $removed_exon (@removed_exons) {
my $locstr = $self->_locstr($removed_exon);
my $inferred_exons = $exon_h{$locstr};
delete $exon_h{$locstr};
if ($inferred_exons) {
my %exons_done = ();
foreach my $exon (@$inferred_exons) {
# make sure we don't move stuff twice
next if $exons_done{$exon};
$exons_done{$exon} = 1;
# we need to tranfer any tag-values from the explicit
# exon to the implicit exon
foreach my $tag ($removed_exon->get_all_tags) {
my @vals = $removed_exon->get_tag_values($tag);
if (!$exon->can("add_tag_value")) {
# I'm puzzled as to what should be done here;
# SeqFeatureIs are not necessarily mutable,
# but we know that in practice the implementing
# class is mutable
$self->throw("The SeqFeature object does not ".
"implement add_tag_value()");
}
$exon->add_tag_value($tag, @vals);
}
}
}
else {
# no exons inferred at $locstr
push(@problems,
[1,
"there is a conflict with exons; there was an explicitly ".
"stated exon with location $locstr, yet I cannot generate ".
"this exon from the supplied mRNA locations\n"]);
}
}
# do we have any inferred exons left over, that were not
# covered in the explicit exons?
if (keys %exon_h) {
# TODO - we ignore this problem for now
push(@problems,
[1,
sprintf("There are some inferred exons that are not in the ".
"explicit exon list; they are the exons at locations:\n".
join("\n", keys %exon_h)."\n")]);
}
# report any problems
if (@problems) {
my $thresh = $self->error_threshold;
my @bad_problems = grep {$_->[0] > $thresh} @problems;
if (@bad_problems) {
printf STDERR "PROBLEM:\n";
$self->_write_hier(\@top_sfs);
# TODO - allow more fine grained control over this
$self->{_problems_reported} = 1;
$self->throw(join("\n",
map {"@$_"} @bad_problems));
}
$self->problem(@$_) foreach @problems;
}
}
}
# --- end of inferring exons --
# return new top level features; this can also
# be retrieved via
# $seq->get_SeqFeatures();
# return @top_sfs;
return $seq->get_SeqFeatures;
}
# _split_group_if_disconnected([@sfs])
#
# as well as having the same group_tag, a group should be spatially
# connected. if not, then the group should be split into subgroups.
# this turns out to be necessary in the case of multicopy genes.
# the standard way to represent these is as spatially disconnected
# gene models (usually a 'gene' feature and some kind of RNA feature)
# with the same group tag; the code below will split these into
# seperate groups, one per copy.
sub _split_group_if_disconnected {
my $self = shift;
my $group = shift;
my @sfs = @$group;
my @ranges =
Bio::Range->disconnected_ranges(@sfs);
my @groups;
if (@ranges == 0) {
$self->throw("ASSERTION ERROR");
}
elsif (@ranges == 1) {
# no need to split the group
@groups = ($group);
}
else {
# @ranges > 1
# split the group into disconnected ranges
if ($self->verbose > 0) {
printf STDERR "GROUP PRE-SPLIT:\n";
$self->_write_group($group, $self->group_tag);
}
@groups =
map {
my $range = $_;
[grep {
$_->intersection($range);
} @sfs]
} @ranges;
if ($self->verbose > 0) {
printf STDERR "SPLIT GROUPS:\n";
$self->_write_group($_, $self->group_tag) foreach @groups;
}
}
return @groups;
}
sub _remove_duplicates_from_group {
my $self = shift;
my $group = shift;
# ::: WEIRD BOUNDARY CASE CODE :::
# for some reason, there are some gb records with two gene
# features for one gene; for example, see ATF14F8.gbk
# in the t/data directory
#
# in this case, we get rid of one of the genes
my @genes = grep {$_->primary_tag eq 'gene'} @$group;
if (@genes > 1) {
# OK, if we look at ATF14F8.gbk we see that some genes
# just exist as a single location, some exist as a multisplit location;
#
# eg
# gene 16790..26395
# /gene="F14F8_60"
# ...
# gene complement(join(16790..19855,20136..20912,21378..21497,
# 21654..21876,22204..22400,22527..23158,23335..23448,
# 23538..23938,24175..24536,24604..24715,24889..24984,
# 25114..25171,25257..25329,25544..25589,25900..26018,
# 26300..26395))
# /gene="F14F8_60"
# the former is the 'standard' way of representing the gene in genbank;
# the latter is redundant with the CDS entry. So we shall get rid of
# the latter with the following filter
if ($self->verbose > 0) {
printf STDERR "REMOVING DUPLICATES:\n";
}
@genes =
grep {
my $loc = $_->location;
if ($loc->isa("Bio::Location::SplitLocationI")) {
my @locs = $loc->each_Location;
if (@locs > 1) {
0;
}
else {
1;
}
}
else {
1;
}
} @genes;
if (@genes > 1) {
# OK, that didn't work. Our only resort is to just pick one at random
@genes = ($genes[0]);
}
if (@genes) {
@genes == 1 || $self->throw("ASSERTION ERROR");
@$group =
($genes[0], grep {$_->primary_tag ne 'gene'} @$group);
}
}
# its a dirty job but someone's gotta do it
return;
}
=head2 unflatten_groups
Title : unflatten_groups
Usage :
Function: iterates over groups, calling unflatten_group() [see below]
Example :
Returns : list of Bio::SeqFeatureI objects that are holders
Args : see below
Arguments
-groups: list of list references; inner list is of Bio::SeqFeatureI objects
e.g. ( [$sf1], [$sf2, $sf3, $sf4], [$sf5, ...], ...)
-resolver_method: a CODE reference
see the documentation above for an example of
a subroutine that can be used to resolve hierarchies
within groups.
this is optional - a default subroutine will be used
NOTE: You should not need to call this method, unless you want fine
grained control over how the unflattening process.
=cut
sub unflatten_groups{
my ($self,@args) = @_;
my($groups, $resolver_method, $resolver_tag) =
$self->_rearrange([qw(GROUPS
RESOLVER_METHOD
RESOLVER_TAG
)],
@args);
# this is just a simple wrapper for unflatten_group()
return
map {
$self->unflatten_group(-group=>$_,
-resolver_method=>$resolver_method,
-resolver_tag=>$resolver_tag)
} @$groups;
}
=head2 unflatten_group
Title : unflatten_group
Usage :
Function: nests a group of features into a feature containment hierarchy
Example :
Returns : Bio::SeqFeatureI objects that holds other features
Args : see below
Arguments
-group: reference to list of Bio::SeqFeatureI objects
-resolver_method: a CODE reference
see the documentation above for an example of
a subroutine that can be used to resolve hierarchies
within groups
this is optional - a default subroutine will be used
NOTE: You should not need to call this method, unless you want fine
grained control over how the unflattening process.
=cut
sub unflatten_group{
my ($self,@args) = @_;
my($group, $resolver_method, $resolver_tag) =
$self->_rearrange([qw(GROUP
RESOLVER_METHOD
RESOLVER_TAG
)],
@args);
if ($self->verbose > 0) {
printf STDERR "UNFLATTENING GROUP:\n";
$self->_write_group($group, $self->group_tag);
}
my @sfs = @$group;
# we can safely ignore singletons (e.g. [source])
return $sfs[0] if @sfs == 1;
my $partonomy = $self->partonomy;
# $resolver_method is a reference to a SUB that will resolve
# ambiguous parent/child containment; for example, determining
# which mRNAs go with which CDSs
$resolver_method = $resolver_method || \&_resolve_container_for_sf;
# TAG BASED RESOLVING OF HIERARCHIES
#
# if the user specifies $resolver_tag, then we use this tag
# to pair up ambiguous parents and children;
#
# for example, the CDS feature may have a resolver tag of /derives_from
# which is a 'foreign key' into the /label tag of the mRNA feature
#
# this kind of tag-based resolution is possible for a certain subset
# of genbank records
#
# if no resolver tag is specified, we revert to the normal
# resolver_method
if ($resolver_tag) {
my $backup_resolver_method = $resolver_method;
# closure: $resolver_tag is remembered by this sub
my $sub =
sub {
my ($self, $sf, @possible_container_sfs) = @_;
my @container_sfs = ();
if ($sf->has_tag($resolver_tag)) {
my ($resolver_tagval) = $sf->get_tag_values($resolver_tag);
# if a feature has a resolver_tag (e.g. /derives_from)
# this specifies the /product, /symbol or /label for the
# parent feature
@container_sfs =
grep {
my $match = 0;
$self->_write_sf($_) if $self->verbose > 0;
foreach my $tag (qw(product symbol label)) {
if ($_->has_tag($tag)) {
my @vals =
$_->get_tag_values($tag);
if (grep {$_ eq $resolver_tagval} @vals) {
$match = 1;
last;
}
}
}
$match;
} @possible_container_sfs;
}
else {
return $backup_resolver_method->($sf, @possible_container_sfs);
}
return map {$_=>0} @container_sfs;
};
$resolver_method = $sub;
}
else {
# CONDITION: $resolver_tag is NOT set
$self->throw("assertion error") if $resolver_tag;
}
# we have now set $resolver_method to a subroutine for
# disambiguatimng parent/child relationships. we will
# now build the whole containment hierarchy for this group
# FIND TOP/ROOT SEQFEATURES
#
# find all the features for which there is no
# containing feature type (eg genes)
my @top_sfs =
grep {
!$self->get_container_type($_->primary_tag);
} @sfs;
# CONDITION: there must be at most one root
if (@top_sfs > 1) {
$self->_write_group($group, $self->group_tag);
printf STDERR "TOP SFS:\n";
$self->_write_sf($_) foreach @top_sfs;
$self->throw("multiple top-sfs in group");
}
my $top_sf = $top_sfs[0];
# CREATE INDEX OF SEQFEATURES BY TYPE
my %sfs_by_type = ();
foreach my $sf (@sfs) {
push(@{$sfs_by_type{$sf->primary_tag}}, $sf);
}
# containment index; keyed by child; lookup parent
# note: this index uses the stringified object reference of
# the object as a surrogate lookup key
my %container = (); # child -> parent
# ALGORITHM: build containment graph
#
# find all possible containers for each SF;
# for instance, for a CDS, the possible containers are all
# the mRNAs in the same group. For a mRNA, the possible
# containers are any SFs of type 'gene' (should only be 1).
# (these container-type mappings can be overridden)
#
# contention is resolved by checking coordinates of splice sites
# (this is the default, but can be overridden)
#
# most of the time, there is no problem identifying a unique
# parent for every child; this can be ambiguous when constructing
# CDS to mRNA relationships with lots of alternate splicing
#
# a hash of child->parent relationships is constructed (%container)
# any mappings that need further resolution (eg CDS to mRNA) are
# placed in %unresolved
# %unresolved index
# (keyed by stringified object reference of child seqfeature)
my %unresolved = (); # child -> [parent,score] to be resolved
# index of seqfeatures by their stringified object reference;
# this is essentially a way of 'reviving' an object from its stringified
# reference
# (see NOTE ON USING OBJECTS AS KEYS IN HASHES, below)
my %idxsf = map {$_=>$_} @sfs;
foreach my $sf (@sfs) {
my $type = $sf->primary_tag;
# container type (e.g. the container type for CDS is usually mRNA)
my $container_type =
$self->get_container_type($type);
if ($container_type) {
my @possible_container_sfs =
@{$sfs_by_type{$container_type} || []};
# we now have a list of possible containers
# (eg for a CDS in an alternately spliced gene, this
# would be a list of all the mRNAs for this gene)
if (!@possible_container_sfs) {
# root of hierarchy
}
else {
if (@possible_container_sfs == 1) {
# this is the easy situation, whereby the containment
# hierarchy is unambiguous. this will probably be the
# case if the genbank record has no alternate splicing
# within it
# ONE OPTION ONLY - resolved!
$container{$sf} = $possible_container_sfs[0];
}
else {
# MULTIPLE CONTAINER CHOICES
$self->throw("ASSERTION ERROR") unless @possible_container_sfs > 1;
# push this onto the %unresolved graph, and deal with it
# later
# for now we hardcode things such that the only type
# with ambiguous parents is a CDS; if this is violated,
# it has a weak problem class of '1' so the API user
# can easily set things to ignore these
if ($sf->primary_tag ne 'CDS') {
$self->problem(1,
"multiple container choice for non-CDS; ".
"CDS to mRNA should be the only ".
"relationships requiring resolving",
$sf);
}
# previously we set the SUB $resolver_method
$self->throw("ASSERTION ERROR")
unless $resolver_method;
# $resolver_method will assign scores to
# parent/child combinations; later on we
# will use these scores to find the optimal
# parent/child pairings
# the default $resolver_method uses splice sites to
# score possible parent/child matches
my %container_sfh =
$resolver_method->($self, $sf, @possible_container_sfs);
if (!%container_sfh) {
$self->problem(2,
"no containers possible for SeqFeature of ".
"type: $type; this SF is being placed at ".
"root level",
$sf);
# RESOLVED! (sort of - placed at root/gene level)
$container{$sf} = $top_sf;
# this sort of thing happens if the record is
# badly messed up and there is absolutely no indication
# of where to put the CDS. Perhaps we should just
# place it with a random mRNA?
}
foreach my $jsf (keys %container_sfh) {
# add [score, parent] pairs to the %unresolved
# lookup table/graph
push(@{$unresolved{$sf}},
[$idxsf{$jsf}, $container_sfh{$jsf} || 0]);
}
}
}
}
else {
# CONDITION:
# not container type for $sf->primary_tag
# CONDITION:
# $sf must be a root/top node (eg gene)
}
}
if (0) {
# CODE CURRENTLY DISABLED
# we require a 1:1 mapping between mRNAs and CDSs;
# create artificial duplicates if we can't do this...
if (%unresolved) {
my %childh = map {$_=>1} keys %unresolved;
my %parenth = map {$_->[0]=>1} map {@$_} values %unresolved;
if ($self->verbose > 0) {
printf STDERR "MATCHING %d CHILDREN TO %d PARENTS\n",
scalar(keys %childh), scalar(keys %parenth);
}
# 99.99% of the time in genbank genomic record of structure type 0, we
# see one CDS for every mRNA; one exception is the S Pombe
# genome, which is all CDS, bar a few spurious mRNAs; we have to
# filter out the spurious mRNAs in this case
#
# another strange case is in the mouse genome, NT_078847.1
# for Pcdh13 you will notice there is 4 mRNAs and 5 CDSs.
# most unusual!
# I'm at a loss for a really clever thing to do here. I think the
# best thing is to create duplicate features to preserve the 1:1 mapping
# my $suffix_id = 1;
# while (keys %childh > keys %parenth) {
#
# }
}
}
# DEBUGGING CODE
if ($self->verbose > 0 && scalar(keys %unresolved)) {
printf STDERR "UNRESOLVED PAIRS:\n";
foreach my $childsf (keys %unresolved) {
my @poss = @{$unresolved{$childsf}};
foreach my $p (@poss) {
my $parentsf = $p->[0];
$childsf = $idxsf{$childsf};
my @clabels = ($childsf->get_tagset_values(qw(protein_id label product)), "?");
my @plabels = ($parentsf->get_tagset_values(qw(transcript_id label product)), "?");
printf STDERR
(" PAIR: $clabels[0] => $plabels[0] (of %d)\n",
scalar(@poss));
}
}
} # -- end of verbose
# Now we have to fully resolve the containment hierarchy; remember,
# the graph %container has the fully resolved child->parent links;
#
# the graph %unresolved is keyed by children missing parents; we
# need to put all these orphans in the %container graph
#
# we do this using the scores in %unresolved, with the
# find_best_matches() algorithm
my $unresolved_problem_reported = 0;
if (%unresolved) {
my $new_pairs =
$self->find_best_matches(\%unresolved, []);
if (!$new_pairs) {
my ($g) = $sfs[0]->get_tagset_values($self->group_tag || 'gene');
$self->problem(2,
"Could not resolve hierarchy for $g");
$new_pairs = [];
$unresolved_problem_reported = 1;
}
foreach my $pair (@$new_pairs) {
if ($self->verbose > 0) {
printf STDERR " resolved pair @$pair\n";
}
$container{$pair->[0]} = $pair->[1];
delete $unresolved{$pair->[0]};
}
}
# CONDITION: containment hierarchy resolved
if (%unresolved) {
$self->throw("UNRESOLVED: %unresolved")
unless $unresolved_problem_reported;
}
# make nested SeqFeature hierarchy from @containment_pairs
# ie put child SeqFeatures into parent SeqFeatures
my @top = ();
foreach my $sf (@sfs) {
my $container_sf = $container{$sf};
if ($container_sf) {
# make $sf nested inside $container_sf
# first check if the container spatially contains the containee
if ($container_sf->contains($sf)) {
# add containee
$container_sf->add_SeqFeature($sf);
}
else {
# weird case - the container does NOT spatially
# contain the containee;
# we expand and throw a warning
#
# for an example of this see ZFP91-CNTF dicistronic gene
# in NCBI chrom 11 build 34.3
$self->problem(1,
"Container feature does not spatially contain ".
"subfeature. Perhaps this is a dicistronic gene? ".
"I am expanding the parent feature",
$container_sf,
$sf);
$container_sf->add_SeqFeature($sf, 'EXPAND');
}
}
else {
push(@top, $sf);
}
}
return @top;
} # -- end of unflatten_group
# -------
# A NOTE ON USING OBJECTS AS KEYS IN HASHES (stringified objects)
#
# Often we with to use seqfeatures as keys in a hashtable; because seqfeatures
# in bioperl have no unique ID, we use a surrogate ID in the form of the
# stringified object references - this is just what you get if you say
#
# print "$sf\n";
#
# this is guaranteed to be unique (within a particular perl execution)
#
# often we want to 'revive' the objects used as keys in a hash - once the
# objects are used as keys, remember it is the *strings* used as keys and
# not the object itself, so the object needs to be revived using another
# hashtable that looks like this
#
# %sfidx = map { $_ => $_ } @sfs
#
# -------
# recursively finds the best set of pairings from a matrix of possible pairings
#
# tries to make sure nothing is unpaired
#
# given a matrix of POSSIBLE matches
# (matrix expressed as hash/lookup; keyed by child object; val = [parent, score]
#
#
sub find_best_matches {
my $self = shift;
my $matrix = shift;
my $pairs = shift; # [child,parent] pairs already selected
my $verbose = $self->verbose;
#################################print "I";
if ($verbose > 0) {
printf STDERR "find_best_matches: (/%d)\n", scalar(@$pairs);
}
my %selected_children = map {($_->[0]=>1)} @$pairs;
my %selected_parents = map {($_->[1]=>1)} @$pairs;
# make a copy of the matrix with the portions still to be
# resolved
my %unresolved_parents = ();
my %unresolved =
map {
if ($verbose > 0) {
printf STDERR " $_ : %s\n", join("; ", map {"[@$_]"} @{$matrix->{$_}});
}
if ($selected_children{$_}) {
();
}
else {
my @parents =
grep {
!$selected_parents{$_->[0]}
} @{$matrix->{$_}};
$unresolved_parents{$_} = 1 foreach @parents;
# new parents
($_ => [@parents]);
}
} keys %$matrix;
my @I = keys %unresolved;
return $pairs if !scalar(keys %unresolved_parents);
# NECESSARY CONDITION:
# all possible parents have a child match
return $pairs if !scalar(@I);
# NECESSARY CONDITION:
# all possible children have a parent match
# give those with fewest choices highest priority
@I = sort {
# n possible parents
scalar(@{$unresolved{$a}})
<=>
scalar(@{$unresolved{$b}}) ;
} @I;
my $csf = shift @I;
my @J = @{$unresolved{$csf}}; # array of [parent, score]
# sort by score, highest first
@J =
sort {
$b->[1] <=> $a->[1]
} @J;
# select pair(s) from remaining matrix of possible pairs
# by iterating through possible parents
my $successful_pairs;
foreach my $j (@J) {
my ($psf, $score) = @$j;
# would selecting $csf, $psf as a pair
# remove all choices from another?
my $bad = 0;
foreach my $sf (@I) {
if (!grep {$_->[0] ne $psf} @{$unresolved{$sf}}) {
# $psf was the only parent choice for $sf
$bad = 1;
last;
}
}
if (!$bad) {
my $pair = [$csf, $psf];
my $new_pairs = [@$pairs, $pair];
my $set = $self->find_best_matches($matrix, $new_pairs);
if ($set) {
$successful_pairs = $set;
last;
}
}
}
# success
return $successful_pairs if $successful_pairs;
# fail
return 0;
}
# ----------------------------------------------
# writes a group to stdout
#
# mostly for logging/debugging
# ----------------------------------------------
sub _write_group {
my $self = shift;
my $group = shift;
my $group_tag = shift || 'gene';
my $f = $group->[0];
my $label = '?';
if ($f->has_tag($group_tag)) {
($label) = $f->get_tag_values($group_tag);
}
if( $self->verbose > 0 ) {
printf STDERR (" GROUP [%s]:%s\n",
$label,
join(' ',
map { $_->primary_tag } @$group));
}
}
sub _write_sf {
my $self = shift;
my $sf = shift;
printf STDERR "TYPE:%s\n", $sf->primary_tag;
return;
}
sub _write_sf_detail {
my $self = shift;
my $sf = shift;
printf STDERR "TYPE:%s\n", $sf->primary_tag;
my @locs = $sf->location->each_Location;
printf STDERR " %s,%s [%s]\n", $_->start, $_->end, $_->strand foreach @locs;
return;
}
sub _write_hier {
my $self = shift;
my @sfs = @{shift || []};
my $indent = shift || 0;
if( $self->verbose > 0 ) {
foreach my $sf (@sfs) {
my $label = '?';
if ($sf->has_tag('product')) {
($label) = $sf->get_tag_values('product');
}
printf STDERR "%s%s $label\n", ' ' x $indent, $sf->primary_tag;
my @sub_sfs = $sf->sub_SeqFeature;
$self->_write_hier(\@sub_sfs, $indent+1);
}
}
}
# -----------------------------------------------
#
# returns all possible containers for an SF based
# on splice site coordinates; splice site coords
# must be contained
# -----------------------------------------------
sub _resolve_container_for_sf{
my ($self, $sf, @possible_container_sfs) = @_;
my @coords = $self->_get_splice_coords_for_sf($sf);
my $start = $sf->start;
my $end = $sf->end;
my $splice_uniq_str = "@coords";
my @sf_score_pairs = ();
# a CDS is contained by a mRNA if the locations of the splice
# coordinates are identical
foreach (@possible_container_sfs) {
my @container_coords = $self->_get_splice_coords_for_sf($_);
my $inside =
!$splice_uniq_str ||
index("@container_coords", $splice_uniq_str) > -1;
if ($inside) {
# the container cannot be smaller than the thing contained
if ($_->start > $start || $_->end < $end) {
$inside = 0;
}
}
# SPECIAL CASE FOR /ribosomal_slippage
# See: http://www.ncbi.nlm.nih.gov/collab/FT/
if (!$inside && $sf->has_tag('ribosomal_slippage')) {
if ($self->verbose > 0) {
printf STDERR " Checking for ribosomal_slippage\n";
}
# TODO: rewrite this to match introns;
# each slippage will be a "fake" small CDS exon
my @transcript_splice_sites = @container_coords;
my @cds_splice_sites = @coords;
##printf STDERR "xxTR SSs: @transcript_splice_sites :: %s\n", $_->get_tag_values('product');
##printf STDERR "xxCD SSs: @cds_splice_sites :: %s\n\n", $sf->get_tag_values('product');
# find the the first splice site within the CDS
while (scalar(@transcript_splice_sites) &&
$transcript_splice_sites[0] < $cds_splice_sites[0]) {
shift @transcript_splice_sites;
}
##print STDERR "TR SSs: @transcript_splice_sites\n";
##print STDERR "CD SSs: @cds_splice_sites\n\n";
if (!(scalar(@transcript_splice_sites)) ||
$transcript_splice_sites[0] == $cds_splice_sites[0]) {
# we will now try and align all splice remaining sites in the transcript and CDS;
# any splice site that can't be aligned is assumed to be a ribosomal slippage
my @slips = ();
my $in_exon = 1;
$inside = 1; # innocent until proven guilty..
while (@cds_splice_sites) {
if (!@transcript_splice_sites) {
# ribosomal slippage is after the last transcript splice site
# Example: (NC_00007, isoform 3 of PEG10)
# mRNA join(85682..85903,92646..99007)
# mRNA join(85682..85903,92646..99007)
# CDS join(85899..85903,92646..93825,93825..94994)
# OR: None of the splice sites align;
# may be a single CDS exon with one slippage inside it.
# Example: (NC_00007, isoform 4 of PEG10)
# mRNA join(85637..85892,92646..99007)
# CDS join(92767..93825,93825..94994)
# Yes, this code is repeated below...
my $p1 = shift @cds_splice_sites;
my $p2 = shift @cds_splice_sites;
if ($self->verbose > 0) {
printf STDERR " Found the ribosomal_slippage: $p1..$p2\n";
}
push(@slips, ($p2-$p1)-1);
}
elsif ($cds_splice_sites[0] == $transcript_splice_sites[0]) {
# splice sites align: this is not the slippage
shift @cds_splice_sites;
shift @transcript_splice_sites;
##print STDERR "MATCH\n";
}
else {
# mismatch
if ($cds_splice_sites[0] < $transcript_splice_sites[0]) {
# potential slippage
# v
# ---TTTTTTTTTT----
# ---CCCC--CCCC----
# ^
my $p1 = shift @cds_splice_sites;
my $p2 = shift @cds_splice_sites;
if ($self->verbose > 0) {
printf STDERR " Found the ribosomal_slippage: $p1..$p2\n";
}
push(@slips, ($p2-$p1)-1);
}
else {
# not a potential ribosomal slippage
$inside = 0; # guilty!
##print STDERR "FAIL\n";
last;
}
}
}
if ($inside) {
# TODO: this is currently completely arbitrary. How many ribosomal slippages do we allow?
# perhaps we need some mini-statistical model here....?
if (@slips > 1) {
$inside = 0;
}
# TODO: this is currently completely arbitrary. What is the maximum size of a ribosomal slippage?
# perhaps we need some mini-statistical model here....?
if (grep {$_ > 2} @slips) {
$inside = 0;
}
}
}
else {
# not a ribosomal_slippage, sorry
}
}
if ($self->verbose > 0) {
printf STDERR " Checking containment:[$inside] (@container_coords) IN ($splice_uniq_str)\n";
}
if ($inside) {
# SCORE: matching (ss-scoords+2)/(n-container-ss-coords+2)
my $score =
(scalar(@coords)+2)/(scalar(@container_coords)+2);
push(@sf_score_pairs,
$_=>$score);
}
}
return @sf_score_pairs;
}
sub _get_splice_coords_for_sf {
my $self = shift;
my $sf = shift;
my @locs = $sf->location;
if ($sf->location->isa("Bio::Location::SplitLocationI")) {
@locs = $sf->location->each_Location;
}
# get an ordered list of (start, end) positions
# my @coords =
# map {
# $_->strand > 0 ? ($_->start, $_->end) : ($_->end, $_->start)
# } @locs;
my @coords = map {($_->start, $_->end)} @locs;
# remove first and last leaving only splice sites
pop @coords;
shift @coords;
return @coords;
}
=head2 feature_from_splitloc
Title : feature_from_splitloc
Usage : $unflattener->feature_from_splitloc(-features=>$sfs);
Function:
Example :
Returns :
Args : see below
At this time all this method does is generate exons for mRNA or other RNA features
Arguments:
-feature: a Bio::SeqFeatureI object (that conforms to Bio::FeatureHolderI)
-seq: a Bio::SeqI object that contains Bio::SeqFeatureI objects
-features: an arrayref of Bio::SeqFeatureI object
=cut
sub feature_from_splitloc{
my ($self,@args) = @_;
my($sf, $seq, $sfs) =
$self->_rearrange([qw(FEATURE
SEQ
FEATURES
)],
@args);
my @sfs = (@{$sfs || []});
push(@sfs, $sf) if $sf;
if ($seq) {
$seq->isa("Bio::SeqI") || $self->throw("$seq NOT A SeqI");
@sfs = $seq->get_all_SeqFeatures;
}
my @exons = grep {$_->primary_tag eq 'exon'} @sfs;
if (@exons) {
$self->problem(2,
"There are already exons, so I will not infer exons");
}
# index of features by type+location
my %loc_h = ();
# infer for every feature
foreach my $sf (@sfs) {
$sf->isa("Bio::SeqFeatureI") || $self->throw("$sf NOT A SeqFeatureI");
$sf->isa("Bio::FeatureHolderI") || $self->throw("$sf NOT A FeatureHolderI");
my $type = $sf->primary_tag;
next unless $type eq 'mRNA' or $type =~ /RNA/;
# an mRNA from genbank will have a discontinuous location,
# with each sub-location being equivalent to an exon
my @locs = $sf->location;
if ($sf->location->isa("Bio::Location::SplitLocationI")) {
@locs = $sf->location->each_Location;
}
if (!@locs) {
use Data::Dumper;
print Dumper $sf;
$self->throw("ASSERTION ERROR: sf has no location objects");
}
# make exons from locations
my @subsfs =
map {
my $subsf = Bio::SeqFeature::Generic->new(-location=>$_,
-primary_tag=>'exon');
## Provide seq_id to new feature:
$subsf->seq_id($sf->seq_id) if $sf->seq_id;
$subsf->source_tag($sf->source_tag) if $sf->source_tag;
## Transfer /locus_tag and /gene tag values to inferred
## features. TODO: Perhaps? this should not be done
## indiscriminantly but rather by virtue of the setting
## of group_tag.
foreach my $tag (grep /gene|locus_tag/, $sf->get_all_tags) {
my @vals = $sf->get_tag_values($tag);
$subsf->add_tag_value($tag, @vals);
}
my $locstr = 'exon::'.$self->_locstr($subsf);
# re-use feature if type and location the same
if ($loc_h{$locstr}) {
$subsf = $loc_h{$locstr};
}
else {
$loc_h{$locstr} = $subsf;
}
$subsf;
} @locs;
# PARANOID CHECK
$self->_check_order_is_consistent($sf->location->strand,@subsfs);
#----
$sf->location(Bio::Location::Simple->new());
# we allow the exons to define the boundaries of the transcript
$sf->add_SeqFeature($_, 'EXPAND') foreach @subsfs;
if (!$sf->location->strand) {
# correct weird bioperl bug in previous versions;
# strand was not being set correctly
$sf->location->strand($subsfs[0]->location->strand);
}
}
return;
}
#sub merge_features_with_same_loc {
# my ($self,@args) = @_;
# my($sfs, $seq) =
# $self->_rearrange([qw(FEATURES
# SEQ
# )],
# @args);
# my @sfs = (@$sfs);
# if ($seq) {
# $seq->isa("Bio::SeqI") || $self->throw("$seq NOT A SeqI");
# @sfs = $seq->get_all_SeqFeatures;
# }
# my %loc_h = ();
# foreach my $sf (@sfs) {
# my $type = $sf->primary_tag;
# my $locstr = $self->_locstr($sf);
## $loc_h{$type.$locstr}
# push(@{$exon_h{$self->_locstr($_)}}, $_) foreach @exons;
# }
#}
=head2 infer_mRNA_from_CDS
Title : infer_mRNA_from_CDS
Usage :
Function:
Example :
Returns :
Args :
given a "type 1" containment hierarchy
gene
CDS
exon
this will infer the uniform "type 0" containment hierarchy
gene
mRNA
CDS
exon
all the children of the CDS will be moved to the mRNA
a "type 2" containment hierarchy is mixed type "0" and "1" (for
example, see ftp.ncbi.nih.gov/genomes/Schizosaccharomyces_pombe/)
=cut
sub infer_mRNA_from_CDS{
my ($self,@args) = @_;
my($sf, $seq, $noinfer) =
$self->_rearrange([qw(FEATURE
SEQ
NOINFER
)],
@args);
my @sfs = ($sf);
if ($seq) {
$seq->isa("Bio::SeqI") || $self->throw("$seq NOT A SeqI");
@sfs = $seq->get_all_SeqFeatures;
}
foreach my $sf (@sfs) {
$sf->isa("Bio::SeqFeatureI") || $self->throw("$sf NOT A SeqFeatureI");
$sf->isa("Bio::FeatureHolderI") || $self->throw("$sf NOT A FeatureHolderI");
if ($self->verbose > 0) {
printf STDERR " Checking $sf %s\n", $sf->primary_tag;
}
if ($sf->primary_tag eq 'mRNA') {
$self->problem(2,
"Inferring mRNAs when there are already mRNAs present");
}
my @cdsl = grep {$_->primary_tag eq 'CDS' } $sf->get_SeqFeatures;
if (@cdsl) {
my @children = grep {$_->primary_tag ne 'CDS'} $sf->get_SeqFeatures;
my @mrnas = ();
foreach my $cds (@cdsl) {
if ($self->verbose > 0) {
print " Inferring mRNA from CDS $cds\n";
}
$self->_check_order_is_consistent($cds->location->strand,$cds->location->each_Location);
my $loc = Bio::Location::Split->new;
foreach my $cdsexonloc ($cds->location->each_Location) {
my $subloc =
Bio::Location::Simple->new(-start=>$cdsexonloc->start,
-end=>$cdsexonloc->end,
-strand=>$cdsexonloc->strand);
$loc->add_sub_Location($subloc);
}
if ($noinfer) {
push(@mrnas, $cds);
}
else {
# share the same location
my $mrna =
Bio::SeqFeature::Generic->new(-location=>$loc,
-primary_tag=>'mRNA');
## Provide seq_id to new feature:
$mrna->seq_id($cds->seq_id) if $cds->seq_id;
$mrna->source_tag($cds->source_tag) if $cds->source_tag;
$self->_check_order_is_consistent($mrna->location->strand,$mrna->location->each_Location);
# make the mRNA hold the CDS; no EXPAND option,
# the CDS cannot be wider than the mRNA
$mrna->add_SeqFeature($cds);
# mRNA steals children of CDS
foreach my $subsf ($cds->get_SeqFeatures) {
$mrna->add_SeqFeature($subsf);
}
$cds->remove_SeqFeatures;
push(@mrnas, $mrna);
}
}
# change gene/CDS to gene/mRNA
$sf->remove_SeqFeatures;
$sf->add_SeqFeature($_) foreach (@mrnas, @children);
}
}
return;
}
=head2 remove_types
Title : remove_types
Usage : $unf->remove_types(-seq=>$seq, -types=>["mRNA"]);
Function:
Example :
Returns :
Args :
removes features of a set type
useful for pre-filtering a genbank record; eg to get rid of STSs
also, there is no way to unflatten
ftp.ncbi.nih.gov/genomes/Schizosaccharomyces_pombe/ UNLESS the bogus
mRNAs in these records are removed (or changed to a different type) -
they just confuse things too much
=cut
sub remove_types{
my ($self,@args) = @_;
my($seq, $types) =
$self->_rearrange([qw(
SEQ
TYPES
)],
@args);
$seq->isa("Bio::SeqI") || $self->throw("$seq NOT A SeqI");
my @sfs = $seq->get_all_SeqFeatures;
my %rh = map {$_=>1} @$types;
@sfs = grep {!$rh{$_->primary_tag}} @sfs;
$seq->remove_SeqFeatures;
$seq->add_SeqFeature($_) foreach @sfs;
return;
}
# _check_order_is_consistent($strand,$ranges) RETURNS BOOL
#
# note: the value of this test is moot - there are many valid,
# if unusual cases where it would flag an anomaly. for example
# transpliced genes such as mod(mdg4) in dmel on AE003744, and
# the following spliced gene on NC_001284:
#
# mRNA complement(join(20571..20717,21692..22086,190740..190761,
# 140724..141939,142769..142998))
# /gene="nad5"
# /note="trans-splicing, RNA editing"
# /db_xref="GeneID:814567"
#
# note how the exons are not in order
# this will flag a level-3 warning, the user of this module
# can ignore this and deal appropriately with the resulting
# unordered exons
sub _check_order_is_consistent {
my $self = shift;
my $parent_strand = shift; # this does nothing..?
my @ranges = @_;
return unless @ranges;
my $rangestr =
join(" ",map{sprintf("[%s,%s]",$_->start,$_->end)} @ranges);
my $strand = $ranges[0]->strand;
for (my $i=1; $i<@ranges;$i++) {
if ($ranges[$i]->strand != $strand) {
$self->problem(1,"inconsistent strands. Trans-spliced gene? Range: $rangestr");
return 1;
# mixed ranges - autopass
# some mRNAs have exons on both strands; for
# example, the dmel mod(mdg4) gene which is
# trans-spliced (in actual fact two mRNAs)
}
}
my $pass = 1;
for (my $i=1; $i<@ranges;$i++) {
my $rangeP = $ranges[$i-1];
my $range = $ranges[$i];
if ($rangeP->start > $range->end) {
if ($self->seq->is_circular) {
# see for example NC_006578.gbk
# we make exceptions for circular genomes here.
# see Re: [Gmod-ajax] flatfile-to-json.pl error with GFF
# 2010-07-26
}
else {
# failed - but still get one more chance..
$pass = 0;
$self->problem(2,"Ranges not in correct order. Strange ensembl genbank entry? Range: $rangestr");
last;
}
}
}
if (!$pass) {
# sometimes (eg ensembl flavour genbank files)
# exons on reverse strand listed in reverse order
# eg join(complement(R1),...,complement(Rn))
# where R1 > R2
for (my $i=1; $i<@ranges;$i++) {
my $rangeP = $ranges[$i-1];
my $range = $ranges[$i];
if ($rangeP->end < $range->start) {
$self->problem(3,"inconsistent order. Range: $rangestr");
return 0;
}
}
}
return 1; # pass
}
# PRIVATE METHOD: _locstr($sf)
#
# returns a location string for a feature; just the outer boundaries
sub _locstr {
my $self = shift;
my $sf = shift;
return
sprintf("%d..%d", $sf->start, $sf->end);
}
sub iterate_containment_tree {
my $self = shift;
my $feature_holder = shift;
my $sub = shift;
$sub->($feature_holder);
my @sfs = $feature_holder->get_SeqFeatures;
$self->iterate_containment_tree($_) foreach @sfs;
}
sub find_best_pairs {
my $matrix = shift;
my $size = shift;
my $i = shift || 0;
for (my $j=0; $j < $size; $j++) {
my $score = $matrix->[$i][$j];
if (!defined($score)) {
next;
}
}
}
1;
|