European Genome-Phenome Archive

File Quality

File InformationEGAF00004997318

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.

5 452 7603 471 5852 787 9382 408 3222 159 6611 969 0251 864 5341 773 8711 729 9301 699 2511 691 3451 699 7411 721 5291 759 5801 808 2021 874 9501 967 2972 088 3842 216 7602 372 5932 549 0492 744 2042 967 0873 214 0483 505 8473 844 3264 268 7414 814 0795 538 4856 485 2457 754 5469 420 40611 568 03214 338 83017 816 88622 075 67927 236 71533 293 41540 267 53248 096 50956 665 67265 804 22875 269 22384 795 71694 029 771102 687 580110 390 431116 878 194121 869 563125 272 619126 911 518126 738 290124 732 981121 144 052116 147 705109 961 255102 856 22395 072 29286 929 56678 658 06470 451 87062 563 73755 123 02548 228 28441 927 41436 260 04331 239 70926 831 83823 021 53519 760 16516 959 64514 594 82212 579 18810 890 7529 458 0148 239 0567 202 1866 297 4845 533 0944 872 3554 294 3713 785 3843 335 4482 945 5162 610 3642 313 1012 049 5681 818 9631 616 1411 442 0281 289 5081 158 9911 042 498940 933855 790783 064719 939668 175619 483579 503548 053516 109489 400468 029447 353427 675412 006394 291378 953362 629350 140337 087324 820315 421304 409294 357285 772275 825267 592259 479250 856242 431237 057230 812223 675218 137212 631206 984202 884196 981192 806187 587182 399177 484173 034168 260164 098160 846156 887153 058149 411144 913142 204138 800134 420132 087129 154126 054124 253120 980117 829116 448113 804111 835110 228107 960105 366103 124102 457100 29998 53196 43994 65093 17391 24089 96088 42887 09284 39982 83081 39580 05978 48876 39575 72174 52673 44371 76070 83268 88668 20267 46365 65564 40363 72162 54761 82160 95260 37459 57158 52056 90956 07055 76354 85253 37153 03952 00051 36051 09050 19049 56848 83648 11146 86246 65745 53945 15344 82643 86943 26042 77643 27942 19941 81841 93341 70341 19840 13139 97239 42239 45538 62938 19737 83236 97936 95236 30936 44536 55536 13635 44935 08634 61034 26033 71833 25132 87232 37232 67532 55431 81832 02931 27730 95230 51630 29229 92129 57729 19829 29428 34228 90928 39727 94227 62427 67727 35127 01226 73426 21426 15025 83625 07625 09824 97324 78624 54424 33624 12324 01623 28322 91523 13922 66322 96622 52522 18321 86621 45921 51121 20720 93321 15820 90920 43120 17620 00219 48119 37019 31719 27418 81218 87518 68718 32418 12718 21917 87417 81917 56917 59317 24717 12617 07916 91816 70716 79516 35316 42816 36515 98015 73515 73015 53115 62215 32815 13514 81614 94614 54414 44214 18014 09013 60914 12313 86713 75213 73713 91713 45713 33312 97813 29412 96713 04412 77012 90212 92412 71612 62512 67712 55112 13112 39111 60311 60511 72411 58111 42211 37311 05611 00310 78510 76410 64010 72310 56710 61910 77610 26710 38010 30910 34510 13410 05110 0329 8519 7789 5859 5609 6209 6329 2659 0529 0948 9658 9198 8708 7958 6178 7318 7508 7068 6478 6308 3238 4508 2378 2018 2058 0138 1317 9928 0918 0958 0017 9027 9257 7347 6327 3147 6077 7587 4857 4697 2447 4777 3867 0337 1547 2757 1277 0846 9916 7027 0827 0466 7536 9076 4486 4886 3906 3216 3076 4616 5116 5576 3116 4146 4146 1896 3036 1866 0116 1366 0475 8455 9885 9386 0766 0465 9605 8855 8725 9385 7285 8345 8935 6695 6355 8175 7695 6695 5955 6475 5375 5075 4945 5355 4405 5735 5885 4355 3035 2865 1915 4065 3605 2345 1865 0914 9504 8835 0614 8765 0225 1795 0525 2795 0274 9724 9824 6524 6824 6234 6584 8764 6354 7614 7814 8054 7274 8114 6104 6054 4994 6314 5944 5074 5174 4124 6304 5214 3824 3254 2874 1504 3304 2134 2884 2964 3404 1574 0334 1454 3594 1324 1603 9904 0554 2484 1534 0104 0213 8594 1024 1013 9493 9183 8633 7403 9213 7903 7633 7353 6343 5353 5023 5663 6163 6763 5563 5273 5683 6053 5303 5053 4893 5483 4133 2913 4523 4283 5513 5633 3673 2963 3183 2663 2213 2383 2273 2413 1813 2493 1573 2693 4173 1713 2163 1883 1983 1783 1763 1993 3503 1093 0553 0523 0363 0623 0762 9722 9842 9133 0973 0202 9372 9012 8732 9263 0412 9943 0562 9662 9852 9252 9102 9542 8752 8882 7842 8912 9172 9922 7902 8432 9012 9232 8832 9842 9272 8342 8712 9422 9762 8662 7762 6832 7682 7842 7092 6532 7182 6032 7102 6562 7702 7532 7192 5272 5042 5272 5702 4012 4182 4692 5142 4932 5112 5652 6412 6452 5462 4252 5532 4572 5802 5752 4382 5372 4992 3482 3302 3472 3232 3012 3522 3472 3232 3952 4012 2942 3682 2942 3242 3332 2392 3702 1982 2662 2832 2022 2902 1932 2732 1592 1112 1672 1592 0612 2742 1132 1742 0602 0932 0432 1682 0732 0292 0262 0881 9672 0902 0122 0652 0781 9431 8381 9211 9421 9171 9462 0021 9841 8871 8951 8802 0011 9561 8691 9531 9581 9191 9411 8832 0472 0052 0761 9411 9651 8031 8892 0391 9691 8931 8441 8821 8321 8831 9501 7821 8601 8171 7911 9131 8581 8201 8451 7921 6431 8701 7631 8681 8961 8631 7731 8161 9071 7821 7581 6711 7491 6911 7451 7481 6941 5801 7471 6581 7551 7131 7661 9001 7091 7831 7761 7251 7111 6751 6801 6051 6741 6671 6261 5361 5821 4631 4581 4831 5251 4681 5571 5761 6111 5901 6121 4601 4891 4661 5431 5071 5471 4831 4921 5211 4231 5751 4231 5031 5251 4831 5131 4821 5201 5541 4801 5241 4361 4561 5181 5231 4541 4521 3731 4441 3811 4051 3671 2851 3391 4751 3511 4061 3471 3981 4081 4161 3951 5021 4931 3541 3381 3341 3891 3561 3661 4011 4581 3151 4311 3551 4081 3791 3761 3661 4291 3881 4561 3551 3731 3091 2621 3991 3661 3551 2631 2921 2531 2531 2571 2361 3191 3081 2271 2691 3191 2711 2251 1631 1461 3291 2871 2081 2581 2831 2091 3151 2611 2331 1991 2461 1971 2331 2251 1771 2081 2441 2101 2121 1921 2011 2051 1731 2411 1991 2161 2101 2361 2741 2031 1731 1421 1861 1171 1501 1591 1081 0881 1041 1621 1341 1551 1601 1851 2121 1041 1371 1211 1611 1541 1061 0281 2031 1211 0651 1231 1001 1521 1891 1161 0971 0551 0321 0141 1161 0971 0791 1301 1671 0211 1561 0421 1151 0211 0089881 0101 0671 0711 0421 0291 0651 0341 0181 0741 0219911 0681 0191 1391 0619791 0799761 0941 0071 0291 0471 0501 0881 0191 0401 0451 0001 0871 0481 1171 0941 0741 0751 0771 0471 0901 0621 0171 0169221 0891 0721 0161 0551 0181 0671 0141 0701 0281 0051 1091 127 200100200300400500600700800900>1000Coverage value1k2k10k20k100k200k1M2M10M20M100M# 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.

1 874 17400000000006 751 358 00200000000000009 972 356 33200000000000143 626 025 71200000510152025303540Phred quality score0G20G40G60G80G100G120G140G# 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).

99.7 %1 058 652 06499.7 %0.3 %

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.

99.5 %1 057 063 54899.5 %0.5 %

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 %1 588 5160.2 %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 %530 965 61050 %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.

97.5 %1 035 091 43297.5 %2.5 %

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.

4.6 %48 448 3844.6 %95.4 %

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.

56 857 6261 347 4631 119 0661 579 3091 282 0531 280 2051 362 5741 649 4401 109 1181 056 159602 626523 060675 984733 110620 699996 159761 299802 491835 307974 1511 021 4641 094 1641 384 6741 023 0071 450 1572 062 915241 8374 308 567290 029267 317531 674455 916284 589590 213258 172240 056362 702428 394171 492705 75012 022 294615 013520 825958 567784 5341 357 1571 436 2291 397 2712 664 957312 121458 141382 314507 594283 803505 067492 059359 5181 301 898336 402683 704955 909 422051015202530354045505560Phred quality score100M200M300M400M500M600M700M800M900M# 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.

99.86%99.84%99.86%99.87%99.86%99.86%99.85%99.85%99.85%99.85%99.85%99.86%99.87%99.85%99.85%99.85%99.84%99.86%99.82%99.84%99.82%99.85%99.67%99.51%0.14%0.16%0.14%0.13%0.14%0.14%0.15%0.15%0.15%0.15%0.15%0.14%0.13%0.15%0.15%0.15%0.16%0.14%0.18%0.16%0.18%0.15%0.33%0.49%123456789101112131415161718192021XYM0%10%20%30%40%50%60%70%80%90%100%mappedunmapped