European Genome-Phenome Archive

File Quality

File InformationEGAF00000642654

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.

561 214 955173 811 79451 921 68519 944 7209 318 5395 678 3704 099 2573 298 2362 828 1362 513 1482 286 6142 109 6801 969 4191 849 9491 753 0821 665 8341 591 5331 517 7001 454 2311 396 0761 345 6641 298 3001 259 4641 220 9021 187 4481 152 6601 120 7091 089 5521 058 3361 031 4301 007 573984 968962 113936 910914 315896 212874 228856 755842 460823 394803 720784 732767 866749 924737 016725 883711 026697 397685 815670 668658 986648 419634 615624 280611 858599 826589 335578 946565 850557 724547 972536 634527 663518 091508 166499 069490 134480 394472 121464 421455 894447 058437 792430 997423 330415 069406 724400 394392 430386 094378 354371 319363 198357 106349 651343 655337 202330 374325 128318 761312 181307 949302 260296 553291 633286 463281 423275 932271 845267 263261 786257 523253 318248 727244 094239 329234 731231 228226 017222 713219 930215 263211 161208 663204 580200 699197 371194 028190 156187 451183 174180 481177 760174 400171 168168 523166 560162 738161 415158 003154 623152 610149 084146 414144 331142 019139 481137 844135 818133 910130 686128 291125 532123 771122 118119 804118 016116 052113 473112 765110 774109 440107 588106 068104 349103 039100 947100 13498 12396 45094 83793 04892 23990 61589 31787 56486 06984 36683 77581 19980 06979 31978 02376 05574 66374 66873 05472 44871 19470 29968 78367 72566 59865 37464 97463 85062 80761 87961 42660 37559 32858 43957 78456 78056 34555 17354 27753 70853 18252 26651 70650 51150 04749 22048 98048 06547 21446 58445 59345 27244 37844 30443 75542 93542 23841 44441 43340 49340 19439 61738 99638 50437 94537 17937 13136 53635 88735 73735 54935 11634 48234 13133 34132 89932 22631 70031 64131 14131 03030 60530 59429 89729 75329 03429 07528 56628 38528 09527 74627 57026 81126 52326 54326 14625 90525 33125 10724 84224 46424 07523 71923 37823 19622 82822 71322 10022 10121 79121 65521 47221 02320 81520 71920 36720 31320 17319 67419 27719 11518 75918 57018 18217 87717 84017 49017 40317 20316 94116 89416 62416 41116 28016 31515 99915 94715 86915 64315 29715 32514 82514 84114 54314 50914 00014 24413 90513 89213 50913 68413 48813 33613 06012 99012 92312 67412 58812 32112 13812 03211 98811 84411 78211 41511 47711 28911 12011 20610 84710 98310 92710 68910 73110 51310 32510 28810 20610 1309 9379 8469 7929 6359 6749 5099 2929 4609 1079 0159 2718 9178 8618 7198 6148 7538 7288 5308 3118 3758 2608 3048 2768 0427 9407 7737 8977 7057 4817 7007 5587 4057 2907 3097 2766 9906 9826 7686 9056 9236 7266 5956 5046 6566 5026 4006 5896 5136 4666 5296 3656 3756 3066 1036 3026 1626 0155 8985 7625 8245 6785 7195 7475 5275 4655 5465 4675 4395 3935 4525 2845 1835 2585 1275 0934 9834 8364 8765 0304 9324 8924 7994 7914 6854 6484 5734 6784 5614 5884 4384 4844 4634 3564 2504 3214 2074 2824 2044 2274 1364 1714 0024 0533 9893 8054 0194 0033 9693 7633 9423 7863 8263 7913 7733 8763 7593 7073 6403 6333 5323 4463 5433 6463 5423 4713 5483 4213 4803 5533 3523 4493 3813 2993 4473 3573 3833 1833 1623 1833 1393 1453 1213 0543 0873 0423 0863 0402 9892 9802 9892 9993 0232 9742 8672 8712 8602 8412 7562 7842 7302 7542 7332 6962 6912 6052 7462 5972 7382 7012 7082 6512 6112 5782 5492 4662 5862 5402 5132 4452 4282 2852 4712 3652 3552 3572 3602 3142 2992 3512 3222 2422 2332 2162 2502 2022 2042 2262 2342 2012 1642 2032 1972 0972 1632 1902 0772 1432 0922 0842 0662 1602 0092 0441 9981 9581 9772 0061 9582 0132 0081 9561 8711 9661 8981 9451 8611 8472 0081 8441 8201 8931 8741 8481 8271 8611 7821 8491 7081 6031 7951 7691 6901 6741 7021 7041 6181 6161 6701 5771 5841 5431 6291 5191 5471 5501 5741 5611 5091 6131 5081 5671 4621 5211 4591 4991 4741 4271 4771 4861 4341 4331 4741 4181 3731 3471 4281 3421 3401 3661 3161 3571 3241 3331 3591 3591 3381 2811 3011 3161 3561 2211 3221 2271 3121 2371 1931 2591 1911 2081 1681 1521 2131 1581 1691 1751 1431 1741 1351 1681 1511 1771 1801 1721 1611 1941 1571 1151 1351 1901 1171 1041 1271 0571 0661 1221 0781 0801 0631 0301 0221 0731 0841 0819931 0381 0669669699799629739441 0109899681 02293392694288992288686288489588787588684885287882789382387084883185281584986478685885883276080676085179980182881078378782584582375081476186381080181578585278984880981680277982076574673074976376371373774074269370868167571068664767171869168366670962668765969865462663168067367862761966262866663759961760664665264361261563963760558862164158664057964662561161457258958853758660258459054756657959354255956255056952155651447355050652255853853953554651754053952348351345149346848847749350149246346247645147649046348844348546944046346641743640641841038041944340241338842637939439137841237738236838137235838140337138738137234134437634434934634637236036837134435934034439837435136634633133632436636336435533130632829433330731531830629632530127330330030031027727326430229028929831128428426728226825727725925027529127230227929326826327130025526725826326522225527328826024325127526225626326624523728222326022622324923522822320222822924623224620823120921523522622021867 263100200300400500600700800900>1000Coverage value1k10k100k1M10M100M# 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.

00399 53907 917 6427 371 5569 876 3867 456 95115 091 9789 119 35312 268 70714 085 40612 677 00917 415 90012 686 79112 795 25515 977 61115 885 82819 657 91419 257 53523 063 66526 061 50726 000 65824 827 09132 679 42735 720 59936 165 16751 428 26344 047 35557 639 26768 050 88787 611 55673 343 995165 906 374142 199 065256 986 726439 792 431705 246 662582 655 3964 399 459 9149 296 58400510152025303540Phred quality score0G0.5G1G1.5G2G2.5G3G3.5G4G# 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).

99.1 %99 090 56299.1 %0.9 %

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.

98.8 %98 793 33698.8 %1.2 %

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 %297 2260.3 %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 %49 987 49350 %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.

98.7 %98 626 33698.7 %1.3 %

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.

5.2 %5 190 4035.2 %94.8 %

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.

9 895 0558 0267 34215 1367 65918 57316 97134 45429 54094 70683 68422 711184 65630 45030 587428 61265 193192 858141 04557 111293 8323 214200 482677 9832 45545 5013 3203 3133 4573 246 4289 1757 1729 58412 31612 63217 892407 5291 186 06634 1308 84063 34434 3424 09694 9844 3649 514274 4669 67620 89414 19034 57616 51453 14450 37080 636148 054361 44081 150 692051015202530354045505560Phred quality score10M20M30M40M50M60M70M80M# 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.

99.75%99.76%99.79%99.62%99.76%99.81%99.66%99.64%99.32%99.54%99.85%99.89%99.82%99.83%99.41%99.45%99.7%99.71%99.77%99.84%99.6%99.69%94.49%99.71%0.25%0.24%0.21%0.38%0.24%0.19%0.34%0.36%0.68%0.46%0.15%0.11%0.18%0.17%0.59%0.55%0.3%0.29%0.23%0.16%0.4%0.31%5.51%0.29%123456789101112131415161718192021XYM0%10%20%30%40%50%60%70%80%90%100%mappedunmapped