European Genome-Phenome Archive

File Quality

File InformationEGAF00000660591

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.

351 405 29771 234 81116 623 3557 923 3765 252 6294 276 1183 697 3533 320 7623 031 0382 810 4392 622 2652 471 1862 335 1282 223 9832 123 5732 038 3931 953 9691 868 9001 797 8731 736 1201 671 6041 604 8031 544 3181 484 2001 426 5691 371 1251 319 6951 265 7881 219 0571 169 0611 120 7401 072 2641 025 149979 238937 064899 097860 605819 079783 393752 299718 161689 155657 709629 808603 708576 953550 381528 239504 852482 871461 955441 570423 530404 507388 398373 230358 209342 905326 736313 376302 662290 565279 587269 219257 216247 107239 026229 620221 166213 670205 828197 938191 292185 555179 305172 256165 168160 237155 254149 595144 472139 241135 234130 111126 481121 628118 244115 216110 681107 124104 183100 99597 93394 03591 71789 00386 09283 62980 58877 72675 03172 99470 73868 26966 60564 85363 20261 15759 26057 91156 21954 46052 50551 73850 25248 44847 05445 79845 22643 24642 22341 47040 33239 59038 27137 21736 73335 19634 50133 70632 33231 37730 91429 80529 14128 64627 62227 10326 49926 12025 29324 48624 27123 57322 83622 31021 80021 40020 84820 37519 75119 17018 69018 25818 04317 81217 11816 65616 49216 04215 75915 27414 73014 44314 26913 86313 52913 18613 01612 93812 50112 08611 86911 74111 27611 39210 94410 76210 47210 19310 1469 8619 5319 3029 0248 9568 6358 4468 3328 1998 1378 0627 8207 7067 5867 5447 3497 2387 1426 7766 7636 6936 4796 4216 4316 1156 0245 7865 8435 6685 6345 4135 4685 2315 0684 9294 8744 9354 7794 7164 6104 4374 3864 3374 3324 2174 2814 1244 1293 9523 8473 9233 9293 7193 7743 6203 7103 6093 6073 3983 4033 3433 3433 2103 1553 0152 8902 8992 8782 8552 8332 7522 6032 7732 8082 5892 4992 5482 4882 4412 3882 3362 3752 3012 3522 2042 1572 2162 1852 1612 0352 1102 1282 1152 0651 9762 0091 9561 9691 9301 9121 9251 8551 8011 8051 7801 7381 6341 7291 6311 6441 5841 5771 5531 4901 4701 5061 3971 4381 3481 4011 3791 3931 3001 2871 3051 3461 2841 2911 2431 2701 1841 1771 2281 2351 2381 2021 2631 2081 1781 2571 2161 1621 1971 1681 1461 1231 1411 1481 1121 0791 0221 0661 0001 0831 1031 0471 0601 0331 0111 0491 0281 0491 0501 0371 0561 0471 04693597093394893794794595092798594489493684987483583081174582583485681080980479377478282678277773676477672878174974072773469172973172373671571370369966466763864368064066466159859869162464162257361564258961561264060160559057164458762959855356157756656552553855151154152851352650548448051949050650248249947948148548247849343241041444442542240940343342240643844845743943645242241941941443241341241643641040039943243541040336839440239237539043039637739736338339035236339234231735737933133834536132729233330831329629333332229835231531229830834532732132834335332234230031128232728430926526630026726829129428827925828327829327823926926224126628628527523023927523425922923722827023426524423824125324826026123024925824923124623724624124923425423321923623422822123021320323222620521719520820021320721121122819418417418917220118519718019017518118318817718119216816317216615017014715917215114016117415813614713414815116016313115114415913912816412912713413512914414614513411012111313911912110612513414413311711211410912311811610410410996112108104961259092939394999298101769110389949510810498988575868193847774777373817677867261675167747557707763686160715968667371687071785557556856615562405074624751515664647170676558615059586262545856584269466354525155445546515661515750525653414651505157464445514647295043514750464046444550494338354144405158442834384729384140443531344944253137363041282444332129374242344441324148292732372533383932383035343341363738283937313438313240524232312433253226264224242926273423253930314329232623212820262224212332332827302230132624252516312929232919211622202435252933232633242713262232212522202328141321242325261314171613151412219161816178 344100200300400500600700800900>1000Coverage value101001k10k100k1M10M100M# 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.

00187 099003 224 6152 700 38411 410 39610 285 0664 526 7616 623 1942 639 6281 911 9954 320 3931 813 5514 724 1284 268 3484 591 7648 505 4355 105 5646 182 3444 276 9037 635 93711 234 97212 594 21414 742 01023 696 63121 770 90620 274 21822 650 15649 107 34281 626 39848 193 39283 825 043168 776 106196 347 742120 772 419318 346 273243 079 009387 035 971462 758 421890 813 92200510152025303540Phred quality score0M100M200M300M400M500M600M700M800M# 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).

98.9 %43 166 18498.9 %1.1 %

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.

98.6 %43 024 09298.6 %1.4 %

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%).

0.3 %142 0920.3 %99.7 %

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 %21 817 19150 %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.

98.2 %42 841 52298.2 %1.8 %

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.

3.2 %1 385 1903.2 %96.8 %

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.

4 890 6355 2653 9399 0174 53812 1449 97519 71617 20750 43239 54013 37782 45217 19114 790179 50134 48991 49668 30224 281131 2041 70379 123287 5111 51918 0072 0132 1821 9851 161 4254 8893 9544 8746 5366 4629 270178 580636 48017 0464 76229 04215 9983 12642 7323 2565 198115 1845 9709 3868 58018 6728 73229 67226 74441 52473 222170 95034 878 582051015202530354045505560Phred quality score5M10M15M20M25M30M# 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.

99.67%99.64%99.78%99.33%99.76%99.82%99.61%99.7%99.42%99.4%99.8%99.84%99.82%99.8%99.6%99.59%99.69%99.64%99.78%99.82%99.56%99.7%95.1%99.82%0.33%0.36%0.22%0.67%0.24%0.18%0.39%0.3%0.58%0.6%0.2%0.16%0.18%0.2%0.4%0.41%0.31%0.36%0.22%0.18%0.44%0.3%4.9%0.18%123456789101112131415161718192021XYM0%10%20%30%40%50%60%70%80%90%100%mappedunmapped