Analysis of the second batch of NimbleGen capture data[edit]
Quick assessment of the data quality[edit]
- First I checked the performance of target capture.
/home/kunzhang/CpgMIP/MONOD/Data/141112_HiSeqRapidRun/get_capture_stats.pl /media/Ext12T/DD_Ext12T/MONOD/141216_HiSeqRapidRun/BAMfiles > 141216_HiSeqRapidRun.capture_stats.txt
Sample
|
total_bases_coverred
|
total_bases
|
on_target_bases_covered
|
on_target_bases
|
Specificity
|
Enrichment_factor
|
6-P-1 |
496,391,712 |
883,938,352 |
20,732,014 |
236,943,256 |
26.81% |
37
|
6-P-2 |
598,767,372 |
974,626,668 |
31,706,660 |
246,684,036 |
25.31% |
34
|
6-P-3 |
543,413,415 |
1,131,193,408 |
31,995,541 |
417,353,349 |
36.89% |
59
|
6T-1 |
729,794,959 |
1,070,341,509 |
30,151,406 |
155,220,183 |
14.50% |
17
|
6T-2 |
784,299,649 |
1,158,440,912 |
29,456,650 |
159,710,882 |
13.79% |
16
|
6T-3 |
1,452,467,042 |
3,323,804,083 |
33,406,665 |
627,923,179 |
18.89% |
24
|
6T-4 |
764,174,440 |
1,132,324,550 |
30,925,708 |
176,622,088 |
15.60% |
19
|
6T-5 |
843,555,327 |
1,282,158,923 |
30,892,326 |
198,472,950 |
15.48% |
19
|
7-P-10 |
465,462,705 |
897,601,881 |
10,665,677 |
208,435,905 |
23.22% |
31
|
7-P-3 |
376,857,145 |
703,513,104 |
10,510,561 |
173,576,316 |
24.67% |
33
|
7-P-5 |
536,571,852 |
807,646,898 |
30,738,662 |
181,239,774 |
22.44% |
29
|
7-P-6 |
616,774,497 |
1,108,444,523 |
20,513,185 |
265,799,682 |
23.98% |
32
|
7-P-7 |
658,461,220 |
1,111,809,575 |
20,479,249 |
228,609,307 |
20.56% |
26
|
7-P-8 |
439,982,190 |
839,669,742 |
11,446,574 |
201,598,027 |
24.01% |
32
|
7T-1 |
780,242,147 |
1,137,145,477 |
30,063,836 |
160,294,067 |
14.10% |
17
|
7T-2 |
746,306,547 |
1,175,101,409 |
26,503,015 |
173,809,990 |
14.79% |
18
|
7T-4 |
771,225,069 |
1,142,595,127 |
29,851,920 |
171,599,332 |
15.02% |
18
|
7T-5 |
802,376,647 |
1,227,075,648 |
27,971,718 |
179,541,336 |
14.63% |
17
|
NC-7 |
255,231,717 |
1,119,499,850 |
7,739,071 |
425,057,131 |
37.97% |
62
|
PC-P_2 |
563,439,590 |
865,434,623 |
30,337,168 |
193,164,906 |
22.32% |
29
|
PC-P_4 |
527,669,028 |
840,929,316 |
29,831,801 |
207,852,537 |
24.72% |
33
|
PC-P_9 |
559,749,434 |
1,035,542,788 |
24,069,681 |
289,633,563 |
27.97% |
39
|
PCT-1 |
714,410,318 |
1,064,633,702 |
27,058,939 |
155,196,610 |
14.58% |
17
|
PCT-2 |
726,354,565 |
1,275,756,897 |
19,580,892 |
189,203,593 |
14.83% |
18
|
PCT-4 |
815,323,371 |
1,247,319,961 |
28,462,046 |
179,280,115 |
14.37% |
17
|
PCT-6 |
948,428,968 |
1,450,561,748 |
31,894,490 |
214,220,998 |
14.77% |
18
|
PCT-7 |
796,647,247 |
1,191,900,342 |
31,314,595 |
191,113,213 |
16.03% |
19
|
|
- Here is the results for the first capture experiment (141112_SN1001).
Sample
|
total_bases_coverred
|
total_bases
|
on_target_bases_covered
|
on_target_bases
|
Specificity
|
Enrichment_factor
|
SeqCap-pCancerSetA-Nov4_12 |
138,931,528 |
1,168,877,279 |
5,883,919 |
450,558,243 |
38.5% |
63
|
SeqCap-pCancerSetA-Nov4_13 |
369,134,874 |
1,424,314,151 |
21,209,194 |
614,835,055 |
43.2% |
77
|
SeqCap-pCancerSetA-Nov4_14 |
302,700,391 |
1,203,488,680 |
13,952,014 |
489,036,528 |
40.6% |
69
|
SeqCap-pCancerSetA-Nov4_15 |
277,562,818 |
1,250,329,814 |
12,577,032 |
511,943,961 |
40.9% |
70
|
SeqCap-pCancerSetA-Nov4_16 |
274,256,170 |
1,164,394,023 |
12,418,863 |
474,550,198 |
40.8% |
70
|
SeqCap-pCancerSetA-Nov4_18 |
403,128,602 |
1,170,296,929 |
23,073,184 |
461,126,140 |
39.4% |
66
|
SeqCap-pCancerSetA-Nov4_19 |
385,167,143 |
1,417,434,873 |
22,614,122 |
614,480,251 |
43.4% |
77
|
SeqCap-pCancerSetA-Nov4_2 |
354,784,810 |
1,514,708,612 |
19,490,111 |
751,025,646 |
49.6% |
99
|
SeqCap-pCancerSetA-Nov4_4 |
352,108,468 |
2,000,885,721 |
23,609,719 |
1,141,652,594 |
57.1% |
134
|
SeqCap-pCancerSetA-Nov4_5 |
196,108,966 |
1,300,234,129 |
8,246,274 |
563,004,318 |
43.3% |
77
|
SeqCap-pCancerSetA-Nov4_6 |
450,502,450 |
1,502,032,451 |
28,135,196 |
645,300,745 |
43.0% |
76
|
SeqCap-pCancerSetA-Nov4_7 |
226,755,475 |
1,432,933,860 |
10,016,683 |
629,536,922 |
43.9% |
79
|
SeqCap-pNormalSetA-Nov4_12 |
368,272,742 |
1,450,833,379 |
32,791,421 |
853,947,465 |
58.9% |
145
|
SeqCap-pNormalSetA-Nov4_13 |
330,611,335 |
1,291,478,405 |
29,764,564 |
756,289,960 |
58.6% |
143
|
SeqCap-pNormalSetA-Nov4_14 |
360,000,998 |
1,364,998,815 |
32,082,346 |
801,469,734 |
58.7% |
144
|
SeqCap-pNormalSetA-Nov4_15 |
375,193,438 |
1,970,841,697 |
30,241,737 |
1,295,408,500 |
65.7% |
194
|
SeqCap-pNormalSetA-Nov4_16 |
365,138,307 |
1,417,821,628 |
31,787,247 |
825,520,233 |
58.2% |
141
|
SeqCap-pNormalSetA-Nov4_18 |
326,952,891 |
1,305,603,890 |
28,937,598 |
760,701,025 |
58.3% |
141
|
SeqCap-pNormalSetA-Nov4_19 |
312,924,926 |
1,271,134,775 |
27,505,559 |
749,645,411 |
59.0% |
145
|
SeqCap-pNormalSetA-Nov4_2 |
353,552,883 |
1,275,196,941 |
31,331,422 |
731,603,678 |
57.4% |
136
|
SeqCap-pNormalSetA-Nov4_4 |
354,621,865 |
1,456,339,179 |
31,829,571 |
867,560,025 |
59.6% |
149
|
SeqCap-pNormalSetA-Nov4_5 |
291,891,632 |
1,066,253,299 |
27,061,797 |
618,610,388 |
58.0% |
140
|
SeqCap-pNormalSetA-Nov4_6 |
333,830,242 |
1,234,362,498 |
31,357,436 |
723,278,371 |
58.6% |
143
|
SeqCap-pNormalSetA-Nov4_7 |
377,475,888 |
1,451,628,039 |
32,325,421 |
843,343,133 |
58.1% |
140
|
|
- For some reason the specificity and enrichment factors for the second experiment is much worse than the first one.
Further comparison of the two batches of library preparation and capturing experiments.[edit]
- There were a number of differences on the experimental protocols between the two batches of data.
- I wrote a script to collect summary statistics for all data sets.
./gather_summary_statistics.pl WGBS_UMR_seqCap_sample_file_info.txt > WGBS_UMR_seqCap_2014_two_batches_summary_stats.txt
- Two major conclusions:
- Kapa Hyper Prep kit generated libraries of higher complexity and lower clonal rates.
- The NimbleGen capture experiment did on 12/15 was not optimal, which lead to much lower enrichment and on-target capture rate.
- Action item: repeat the capture on the batch of libraries prepared by the Kapa Hyper Prep kit.
Sample
|
Experiment
|
N_mapped_reads
|
N_non-clonal_reads
|
N_on-target_reads
|
N_non-clonal_on-target_reads
|
N_on-target_haplotypes
|
N_target_coverred
|
Pct_on-target
|
Pct_clonal
|
6-P-1 |
141112_HiSeq |
14,727,691 |
12,016,120 |
6,119,021 |
4,477,865 |
336,894 |
38,344 |
41.5% |
26.82%
|
6-P-1 |
141216_HiSeq |
10,355,738 |
10,123,240 |
2,619,216 |
2,523,753 |
233,941 |
36,713 |
25.3% |
3.64%
|
6-P-10 |
141112_HiSeq |
12,125,750 |
10,627,443 |
4,622,479 |
3,674,457 |
389,005 |
39,594 |
38.1% |
20.51%
|
6-P-2 |
141112_HiSeq |
12,610,235 |
10,592,101 |
4,890,268 |
3,776,213 |
194,655 |
30,935 |
38.8% |
22.78%
|
6-P-2 |
141216_HiSeq |
11,570,287 |
11,462,431 |
2,756,445 |
2,724,126 |
882,786 |
44,945 |
23.8% |
1.17%
|
6-P-3 |
141112_HiSeq |
13,044,553 |
10,902,745 |
5,089,409 |
3,951,200 |
180,681 |
29,498 |
39.0% |
22.36%
|
6-P-3 |
141216_HiSeq |
12,482,525 |
12,258,142 |
4,304,444 |
4,228,174 |
1,154,014 |
45,181 |
34.5% |
1.77%
|
6-P-4 |
141112_HiSeq |
11,969,782 |
9,696,006 |
4,662,823 |
3,426,251 |
184,637 |
29,454 |
39.0% |
26.52%
|
6-P-8 |
141112_HiSeq |
14,935,803 |
13,198,763 |
6,242,428 |
5,177,911 |
351,452 |
38,772 |
41.8% |
17.05%
|
6T-1 |
141216_HiSeq |
11,869,709 |
11,711,949 |
1,631,628 |
1,592,334 |
614,412 |
42,951 |
13.7% |
2.41%
|
6T-2 |
141216_HiSeq |
12,820,323 |
12,644,365 |
1,679,917 |
1,605,021 |
535,022 |
42,471 |
13.1% |
4.46%
|
6T-3 |
141216_HiSeq |
36,126,640 |
33,993,548 |
6,294,978 |
5,456,336 |
1,390,560 |
45,913 |
17.4% |
13.32%
|
6T-4 |
141216_HiSeq |
12,351,984 |
12,219,697 |
1,829,796 |
1,794,578 |
722,787 |
43,545 |
14.8% |
1.92%
|
6T-5 |
141216_HiSeq |
13,871,915 |
13,700,144 |
2,047,985 |
2,002,834 |
689,818 |
43,576 |
14.8% |
2.20%
|
7-P-10 |
141216_HiSeq |
10,375,497 |
10,108,671 |
2,264,332 |
2,166,347 |
108,343 |
25,759 |
21.8% |
4.33%
|
7-P-2 |
141112_HiSeq |
12,071,295 |
10,045,638 |
4,407,555 |
3,544,280 |
86,126 |
17,973 |
36.5% |
19.59%
|
7-P-3 |
141216_HiSeq |
8,343,291 |
8,131,342 |
1,913,926 |
1,834,202 |
103,372 |
25,099 |
22.9% |
4.17%
|
7-P-5 |
141216_HiSeq |
9,868,792 |
9,790,293 |
2,096,290 |
2,077,736 |
679,672 |
44,364 |
21.2% |
0.89%
|
7-P-6 |
141216_HiSeq |
13,420,765 |
13,142,028 |
3,044,464 |
2,934,292 |
235,624 |
36,563 |
22.7% |
3.62%
|
7-P-7 |
141216_HiSeq |
13,264,972 |
13,020,148 |
2,601,234 |
2,509,390 |
217,498 |
37,099 |
19.6% |
3.53%
|
7-P-8 |
141216_HiSeq |
10,309,480 |
10,046,649 |
2,320,567 |
2,222,588 |
120,025 |
26,514 |
22.5% |
4.22%
|
7T-1 |
141216_HiSeq |
12,579,473 |
12,449,866 |
1,685,639 |
1,644,912 |
593,556 |
43,006 |
13.4% |
2.42%
|
7T-2 |
141216_HiSeq |
13,386,508 |
13,115,388 |
1,875,500 |
1,768,594 |
402,132 |
40,445 |
14.0% |
5.70%
|
7T-4 |
141216_HiSeq |
12,632,367 |
12,486,042 |
1,802,005 |
1,748,412 |
578,586 |
42,950 |
14.3% |
2.97%
|
7T-5 |
141216_HiSeq |
13,465,056 |
13,277,929 |
1,888,156 |
1,831,556 |
440,050 |
41,621 |
14.0% |
3.00%
|
NC-12 |
141112_HiSeq |
13,489,901 |
11,847,524 |
7,775,540 |
6,528,741 |
668,144 |
43,880 |
57.6% |
16.03%
|
NC-13 |
141112_HiSeq |
14,739,045 |
13,565,791 |
8,516,530 |
7,634,190 |
1,026,188 |
45,600 |
57.8% |
10.36%
|
NC-14 |
141112_HiSeq |
20,083,892 |
18,545,178 |
12,972,574 |
11,822,645 |
963,541 |
44,221 |
64.6% |
8.86%
|
NC-15 |
141112_HiSeq |
14,827,402 |
13,268,019 |
8,506,375 |
7,318,388 |
1,009,294 |
45,460 |
57.4% |
13.97%
|
NC-17 |
141112_HiSeq |
13,336,373 |
11,378,302 |
7,670,034 |
6,154,378 |
702,115 |
43,812 |
57.5% |
19.76%
|
NC-18 |
141112_HiSeq |
13,248,090 |
11,294,433 |
7,676,163 |
6,176,274 |
513,023 |
42,743 |
57.9% |
19.54%
|
NC-2 |
141112_HiSeq |
13,866,539 |
12,827,193 |
7,830,036 |
7,062,215 |
881,306 |
45,316 |
56.5% |
9.81%
|
NC-21 |
141112_HiSeq |
16,081,583 |
14,965,703 |
9,245,026 |
8,425,521 |
1,436,801 |
46,192 |
57.5% |
8.86%
|
NC-3 |
141112_HiSeq |
16,027,488 |
14,330,418 |
9,380,987 |
8,102,323 |
1,031,698 |
45,529 |
58.5% |
13.63%
|
NC-5 |
141112_HiSeq |
11,531,790 |
10,057,121 |
6,516,165 |
5,388,840 |
478,433 |
42,318 |
56.5% |
17.30%
|
NC-6 |
141112_HiSeq |
13,434,528 |
12,080,666 |
7,733,596 |
6,691,809 |
867,491 |
45,159 |
57.6% |
13.47%
|
NC-7 |
141112_HiSeq |
15,290,119 |
13,970,372 |
8,748,247 |
7,744,173 |
1,137,553 |
45,946 |
57.2% |
11.48%
|
NC-7 |
141216_HiSeq |
11,685,910 |
9,211,663 |
4,179,919 |
2,929,376 |
122,228 |
22,142 |
35.8% |
29.92%
|
PC-P_2 |
141216_HiSeq |
10,306,946 |
10,206,818 |
2,171,141 |
2,140,765 |
637,877 |
44,379 |
21.1% |
1.40%
|
PC-P_4 |
141216_HiSeq |
10,561,291 |
10,419,994 |
2,476,732 |
2,423,078 |
568,230 |
43,728 |
23.5% |
2.17%
|
PC-P_9 |
141216_HiSeq |
12,272,000 |
11,995,732 |
3,261,610 |
3,143,222 |
316,171 |
39,834 |
26.6% |
3.63%
|
PC-P-3 |
141112_HiSeq |
15,157,128 |
12,617,188 |
7,256,675 |
5,581,424 |
340,207 |
36,891 |
47.9% |
23.09%
|
PC-P-4 |
141112_HiSeq |
19,403,398 |
15,071,020 |
10,744,097 |
7,603,427 |
596,193 |
39,582 |
55.4% |
29.23%
|
PC-P-5 |
141112_HiSeq |
13,196,672 |
10,533,012 |
5,415,343 |
4,051,510 |
131,556 |
22,886 |
41.0% |
25.18%
|
PC-P-6 |
141112_HiSeq |
15,392,120 |
13,176,909 |
6,418,169 |
4,958,048 |
604,369 |
43,138 |
41.7% |
22.75%
|
PC-P-7 |
141112_HiSeq |
14,218,885 |
10,784,660 |
5,965,548 |
4,124,202 |
176,841 |
26,171 |
42.0% |
30.87%
|
PCT-1 |
141216_HiSeq |
11,921,316 |
11,737,650 |
1,655,540 |
1,607,618 |
389,989 |
41,033 |
13.9% |
2.89%
|
PCT-2 |
141216_HiSeq |
14,465,075 |
14,097,386 |
2,031,726 |
1,932,100 |
239,455 |
35,730 |
14.0% |
4.90%
|
PCT-4 |
141216_HiSeq |
13,781,204 |
13,569,716 |
1,895,572 |
1,841,279 |
458,181 |
41,897 |
13.8% |
2.86%
|
PCT-6 |
141216_HiSeq |
15,681,405 |
15,532,475 |
2,186,241 |
2,149,561 |
882,376 |
44,422 |
13.9% |
1.68%
|
PCT-7 |
141216_HiSeq |
12,980,554 |
12,835,501 |
1,973,329 |
1,938,285 |
760,032 |
43,769 |
15.2% |
1.78%
|
|