European Genome-Phenome Archive

File Quality

File InformationEGAF00008064815

File Data

Base Coverage Distribution

This chart represents the base coverage distribution along the reference file. Y-axis represents the number of times a position in the reference file is covered. The x-axis represents the range of the values for the coverage.

Data is represented in a log scale to minimise the variability. A high peak in the beginning (low coverage) and a curve descending is expected.

5 724 4133 173 3962 478 1562 187 2281 994 8821 838 3601 713 0701 612 0791 512 9591 429 9441 377 9721 328 1701 275 7891 231 2721 201 9911 184 8691 160 2801 144 5941 134 8671 132 0511 133 3941 144 5651 157 4081 176 0391 197 5221 232 3521 268 6441 317 7021 369 5791 421 9051 485 0041 561 8831 636 7971 714 2011 781 3071 865 0311 953 4622 039 3862 124 3012 219 8512 326 2032 448 2082 583 1632 760 0472 968 7073 224 6633 555 0023 965 7844 475 0195 118 2205 906 1606 875 0238 052 2419 469 38511 167 81113 157 04615 494 04618 189 10621 227 21024 656 92628 462 03332 633 49737 147 87941 944 20846 961 23052 198 83857 522 19362 831 57068 066 51373 084 17077 811 83682 183 61386 026 90589 319 87391 955 84993 960 85595 161 31495 709 70895 488 66794 537 92592 894 37490 609 50887 737 56984 363 61580 582 92676 441 40872 023 81467 427 33962 728 83858 003 68253 317 93848 753 09544 334 02740 100 44836 113 80732 369 32028 889 83425 678 80722 739 40120 072 81817 651 53415 495 63813 569 25511 855 67810 337 5019 027 6987 861 6546 853 8185 985 8355 233 6444 578 0934 010 4083 525 2053 104 1992 743 1102 434 9292 164 8171 936 3271 736 7731 562 0121 411 9511 283 0931 169 8921 071 782985 228909 150841 681781 224727 807683 825642 984607 527577 169544 702515 411493 092469 244449 789432 060415 145399 817386 825374 504363 407352 461341 761333 447325 219316 111309 806301 782296 194289 117281 852275 045269 980263 040257 651253 064245 865242 085234 203229 269223 877219 254214 174209 259205 003199 370195 119191 698187 559182 689178 664174 392172 291168 930165 078162 172158 710156 063152 507149 236147 101144 115141 588140 072137 468134 117131 985129 599126 910124 670123 509120 836118 434116 107114 704114 178112 930109 970108 896107 115106 691105 351104 017102 407100 16899 03398 19097 01296 36494 82593 91492 95591 08990 23988 59588 17587 56986 29085 56684 20283 32081 77380 73379 77878 90778 38177 50376 29974 48273 82173 14372 09571 51270 25269 49568 42667 51167 27066 34465 04663 94062 64661 78060 65460 44160 13959 13858 00257 23356 97655 65654 57954 49153 71253 07652 48951 54151 32450 86649 65849 43349 17647 94247 59746 74046 51446 00145 32245 09344 81843 78843 09142 51742 23242 13741 23241 00340 59540 52139 92539 23139 13138 59938 16837 65837 14436 91436 60936 09335 51934 74334 81434 23133 76632 99732 82532 58932 62832 70532 27532 01231 96231 38731 19530 62030 29730 21230 14229 99029 67829 18329 02329 16828 60628 34827 97628 50727 82827 53327 04527 04226 91726 36226 49926 18225 36926 11225 69625 30425 27624 99324 96724 55124 76224 52224 41824 22124 14423 90723 71323 48723 14323 33023 21222 96322 91322 97622 57222 48322 33622 20122 06021 89021 63721 66421 58921 70121 48121 43721 20920 90020 89320 85620 48820 33020 31320 10719 93719 61119 75519 89019 57119 86719 46919 57519 64819 31419 09019 13618 89818 96018 65918 40218 67518 18818 07018 12018 04517 95517 74717 62417 52817 25617 24517 10316 91216 61816 34116 34216 56616 31216 22915 90115 81115 97315 82515 53415 47515 38015 29915 21415 13515 05214 73214 63314 86914 70614 71214 60914 46614 61414 62314 17214 36514 10414 02313 80113 54013 81513 64413 57313 31713 47013 40813 27413 11713 14912 84112 99612 77112 89612 70712 73212 41212 72212 17812 35512 26912 17012 26512 17011 86311 82611 70711 72311 35611 61211 46511 26711 37711 13511 44911 23511 36311 22911 23110 87711 23110 69210 87510 77110 65710 45810 45410 27610 29910 41010 00110 26710 21610 0089 8619 95910 1239 8209 8299 6869 6939 7129 2369 5399 5009 4319 1629 1779 1809 0199 0299 1399 0858 9688 8708 9678 8178 8508 6768 8288 8348 5868 6408 4708 2828 3078 4358 1947 9848 0418 0698 0357 7967 8537 8237 7477 6577 8367 6907 5997 5487 5867 3707 4447 5857 4047 5097 4117 3187 2577 1457 4227 0427 0947 1327 1316 8756 9546 9166 9326 7786 7646 8166 6646 7206 6106 7886 4916 5946 7736 5376 5586 4996 3996 3696 4556 2066 2306 2276 2526 1676 2216 1396 0746 1456 2876 0205 8615 9705 8866 0135 6815 8255 8795 9565 8065 9365 8245 8285 7745 7635 7595 7195 8175 7775 5475 6385 5185 4015 3065 5305 2775 3335 4155 4995 1415 1175 1285 2525 0675 0775 2144 9595 0745 1285 0605 0375 1274 9675 0424 9964 9744 8494 9715 0154 8654 7784 6244 7084 6004 7094 6914 5694 7034 6074 5554 4294 4234 4584 4204 3674 4214 3534 2594 3644 3574 2764 1774 1284 2494 2054 3084 2044 3184 1344 2104 1914 2374 1174 1764 1724 0554 1374 0044 0603 9593 8463 9604 0043 9603 8873 9033 9814 0904 0114 0273 9803 9223 9283 8763 8543 8003 8813 8133 7313 8003 6923 7643 8303 6553 7963 6193 6143 6313 6583 6413 7453 6333 7183 4383 5043 5223 6023 5123 5233 4103 5453 4713 5603 3523 3323 3863 2693 2213 2473 3753 2823 4513 3603 3693 1623 1343 3413 2413 2843 2663 2223 2353 3793 2533 3143 1203 1883 1693 2083 0513 1433 1293 1153 1853 0923 1343 1932 9952 9703 1143 0353 1783 1083 0833 0483 0393 0893 0983 0242 9923 0553 0223 1182 9082 9823 0462 8542 9902 9392 9142 9102 9732 8992 8412 8412 8292 8992 7682 9022 8442 7702 8212 8592 7672 7432 7502 8052 8542 7702 6802 7502 7522 7572 7342 6812 7102 7212 7032 6582 6532 6012 6512 6952 5522 6582 5962 5582 5282 5902 6232 4932 4952 5022 5022 4072 4492 3922 4382 3712 4542 4632 4072 3682 3092 3302 3442 3722 3432 3792 3512 4232 2752 3472 3152 3452 3342 3482 3672 2642 3432 3052 2982 2332 2212 2082 2872 2802 2562 3102 1892 2192 1882 2162 1482 1202 2012 1602 1592 1052 2012 0832 0942 2592 1232 1372 2382 1432 0442 0512 0582 1962 1382 1212 1022 1062 1362 1902 1171 9411 9692 0891 9511 8812 0322 0042 0171 9492 0072 0582 0161 9912 0111 9772 0192 0472 0491 9661 9731 9691 9051 9151 8331 8881 9631 9071 9291 9521 9561 9001 8791 8921 8221 9601 9161 7561 9581 8461 8201 8211 7981 8021 7621 7691 7661 8031 7411 8001 7801 7101 6721 7341 7361 6611 6701 7591 6321 7761 7431 6971 6931 7781 6501 6661 6011 6251 6141 6531 6091 6631 5501 4851 5301 5921 5431 5311 5271 6161 5341 5091 6461 4851 5651 6581 5771 5531 5721 5931 6071 5241 4171 5581 5031 4651 5181 4761 4841 4741 4781 4641 4621 4171 4771 5151 5621 5641 4681 5051 4281 3851 4521 4721 4201 3911 4491 3681 3681 4111 5051 3991 3541 3951 4511 4601 3741 535 748100200300400500600700800900>1000Coverage value2k10k20k100k200k1M2M10M20M# Bases

Base Quality

The base quality distribution shows the Phred quality scores describing the probability that a nucleotide has been incorrectly assigned; e.g. an error in the sequencing. Specifically, Q=-log10(P), where Q is the Phred score and P is the probability the nucleotide is wrong. The larger the score, the more confident we are in the base call. Depending on the sequencing technology, we can expect to see different distributions, but we expect to see a distribution skewed towards larger (more confident) scores; typically around 40.

18 790 76700000000007 853 766 686000000000000012 553 047 83200000000000219 052 285 41100000510152025303540Phred quality score0G20G40G60G80G100G120G140G160G180G200G# Bases

Mapped Reads

Number of reads successfully mapped (singletons & both mates) to the reference genome in the sample. Genetic variation, in particular structural variants, ensure that every sequenced sample is genetically different from the reference genome it was aligned to. Small differences against the reference are accepted, but, for more significant variation, the read can fail to be placed. Therefore, it is not expected that the mapped reads rate will hit 100%, but it is supposed to be high (usually >90%). Calculations are made taking into account the proportion of mapped reads against the total number of reads (mapped/mapped+unmapped).

99.8 %1 583 246 33299.8 %0.2 %

Both Mates Mapped

When working with paired-end sequencing, each DNA fragment is sequenced from both ends, creating two mates for each pair. This chart shows the fraction of reads in pairs where both of the mates successfully map to the reference genome. .

Notice that reads not mapped to the expected distance are also included as occurs with the proper pairs chart.

99.7 %1 580 829 36899.7 %0.3 %

Singletons

When working with paired-end sequencing, each DNA fragment is sequenced from both ends, creating two mates for each pair. If one mate in the pair successfully maps to the reference genome, but the other is unmapped, the mapped mate is a singleton. One way in which a singleton could occur would be if the sample has a large insertion compared with the reference genome; one mate can fall in sequence flanking the insertion and will be mapped, but the other falls in the inserted sequence and so cannot map to the reference genome. There are unlikely to many such structural variants in the sample, or sequencing errors that would cause a read not to be able to map. Consequently, the singleton rate is expected to be very low (<1%).

0.2 %2 416 9640.2 %99.8 %

Forward Strand

Fraction of reads mapped to the forward DNA strand. The general expectation is that the DNA library preparation step will generate DNA from the forward and reverse strands in equal amounts so after mapping the reads to the reference genome, approximately 50% of them will consequently map to the forward strand. Deviations from the 50%, may be due to problems with the library preparation step.

50 %792 973 14850 %50 %

Proper Pairs

A fragment consisting of two mates is called a proper pair if both mates map to the reference genome at the expected distance according to the reference genome. In particular, if the DNA library consists of fragments ~500 base pairs in length, and 100 base pair reads are sequenced from either end, the expectation would be that the two reads map to the reference genome separated by ~300 base pairs. If the sequenced sample contains large structural variants, e.g. a large insertion, where we expect the reads mapping with a large separation would be a signal for this variant, and the reads would not be considered as proper pairs. Based on the sequencing technology, there is also an expectation of the orientation of each read in the fragment.

The rate of proper pairs is expected to be well over 90%; even if the mapping rate itself is low as a result of bacterial contamination, for example.

97.3 %1 542 678 25697.3 %2.7 %

Duplicates

PCR duplicates are two (or more) reads that originate from the same DNA fragment. When sequencing data is analyzed, it is assumed that each observation (i.e. each read) is independent; an assumption that fails in the presence of duplicate reads. Typically, algorithms look for reads that map to the same genomic coordinate, and whose mates also map to identical genomic coordinates. It is important to note that as the sequencing depth increases, more reads are sampled from the DNA library, and consequently it is increasingly likely that duplicate reads will be sampled. As a result, the true duplicate rate is not independent of the depth, and they should both be considered when looking at the duplicate rate. Additionally, as the sequencing depth in increases, it is also increasingly likely that reads will map to the same location and be marked as duplicates, even when they are not. As such, as the sequencing depth approaches and surpasses the read length, the duplicate rate starts to become less indicative of problems.

9.4 %148 641 4879.4 %90.6 %

Mapping Quality Distribution

The mapping quality distribution shows the Phred quality scores describing the probability that a read does not map to the location that it has been assigned to (specifically, Q=-log10(P), where Q is the Phred score and P is the probability the read is in the wrong location). So the larger the score, the higher the quality of the mapping. Some scores have a specific meaning, e.g. a score of 0 means that the read could map equally to multiple places in the reference genome. The majority of reads should be well mapped, and so we expect to see this distribution heavily skewed to a significant value (typically around 60). It is not unusual to see some scores around zero. Reads originating from repetitive elements in the genome will plausibly map to multiple locations.

76 545 6591 728 8111 373 5562 068 4091 561 2501 591 0211 718 5832 354 3071 520 9101 373 201753 619655 963874 400986 963812 2561 326 875976 6761 065 6901 060 3751 276 2911 335 3411 443 8901 892 5231 356 3961 932 3852 999 438313 4686 389 787373 826349 649733 945633 732423 348787 986355 250327 368495 833614 479232 0591 009 97217 035 513877 514697 3541 384 7761 096 6001 892 7422 182 0241 839 7034 171 363411 318668 739519 397732 426365 056707 290637 824449 0421 928 146417 073933 2871 438 042 101051015202530354045505560Phred quality score0.2G0.4G0.6G0.8G1G1.2G1.4G# Reads

Mapped vs Unmapped

Stacked column chart for both mapped and unmapped reads along all chromosomes in the reference file. It is a similar representation as shown in the Mapped reads chart but for each chromosome. Although sequenced sample may be a female, it is possible to get reads in the Y chromosome as there are common regions in both chromosomes called pseudoautosomal regions (PAR1, PAR2).

Unmapped reads belonging to each chromosome are determined when the one mate/pair is aligned and the other is not. The unmapped read should have chromosome and POS identical to its mate. It could also be due when aligning is performed with bwa as it concatenates all the reference sequences together, so if a read hangs off of one reference onto another, it will be given the right chromosome and position, but it also be classified as unmapped.

99.85%99.84%99.86%99.87%99.86%99.86%99.85%99.85%99.85%99.84%99.85%99.85%99.87%99.85%99.84%99.84%99.83%99.86%99.81%99.83%99.81%99.85%99.64%99.55%0.15%0.16%0.14%0.13%0.14%0.14%0.15%0.15%0.15%0.16%0.15%0.15%0.13%0.15%0.16%0.16%0.17%0.14%0.19%0.17%0.19%0.15%0.36%0.45%123456789101112131415161718192021XYM0%10%20%30%40%50%60%70%80%90%100%mappedunmapped