Open chromatin assays are known to have significant GC bias. Please take this
into consideration as necessary.
Library complexity quality metrics
Library complexity (filtered non-mito BAM)
rep1
rep2
rep3
rep4
ctl1
ctl2
ctl3
ctl4
Total Fragments
1582659
1521891
588320
1075267
1405680
1919660
2794172
1964730
Distinct Fragments
1540009
1505393
571973
1023368
1402051
1913239
2780126
1958152
Positions with Two Read
38289
15687
15098
46005
3498
6284
13066
6370
NRF = Distinct/Total
0.973052
0.98916
0.972214
0.951734
0.997418
0.996655
0.994973
0.996652
PBC1 = OneRead/Distinct
0.97389
0.989326
0.972591
0.952367
0.997461
0.99668
0.995139
0.996695
PBC2 = OneRead/TwoRead
39.170493
94.940014
36.845675
21.185132
399.797313
303.451305
211.741237
306.386342
Mitochondrial reads are filtered out by default.
The non-redundant fraction (NRF) is the fraction of non-redundant mapped reads
in a dataset; it is the ratio between the number of positions in the genome
that uniquely mapped reads map to and the total number of uniquely mappable
reads. The NRF should be > 0.8. The PBC1 is the ratio of genomic locations
with EXACTLY one read pair over the genomic locations with AT LEAST one read
pair. PBC1 is the primary measure, and the PBC1 should be close to 1.
Provisionally 0-0.5 is severe bottlenecking, 0.5-0.8 is moderate bottlenecking,
0.8-0.9 is mild bottlenecking, and 0.9-1.0 is no bottlenecking. The PBC2 is
the ratio of genomic locations with EXACTLY one read pair over the genomic
locations with EXACTLY two read pairs. The PBC2 should be significantly
greater than 1. See more details at
the ENCODE portal standard for ChIP-Seq pipeline
NRF (non redundant fraction)
PBC1 (PCR Bottleneck coefficient 1)
PBC2 (PCR Bottleneck coefficient 2)
PBC1 is the primary measure. Provisionally
0-0.5 is severe bottlenecking
0.5-0.8 is moderate bottlenecking
0.8-0.9 is mild bottlenecking
0.9-1.0 is no bottlenecking
Replication quality metrics
IDR (Irreproducible Discovery Rate) plots
Reproducibility QC and peak detection statistics
overlap
idr
Nt
1720
833
N1
3607
987
N2
2865
843
N3
3328
776
N4
3604
870
Np
2524
914
N optimal
2524
914
N conservative
1720
833
Optimal Set
pooled-pr1_vs_pooled-pr2
pooled-pr1_vs_pooled-pr2
Conservative Set
rep1_vs_rep2
rep3_vs_rep4
Rescue Ratio
1.4674418604651163
1.0972388955582233
Self Consistency Ratio
1.2589877835951135
1.2719072164948453
Reproducibility Test
pass
pass
Reproducibility QC
N1: Replicate 1 self-consistent peaks (comparing two pseudoreplicates generated by subsampling Rep1 reads)
N2: Replicate 2 self-consistent peaks (comparing two pseudoreplicates generated by subsampling Rep2 reads)
Ni: Replicate i self-consistent peaks (comparing two pseudoreplicates generated by subsampling RepX reads)
Nt: True Replicate consistent peaks (comparing true replicates Rep1 vs Rep2)
Np: Pooled-pseudoreplicate consistent peaks (comparing two pseudoreplicates generated by subsampling pooled reads from Rep1 and Rep2)
Self-consistency Ratio: max(N1,N2) / min (N1,N2)
Rescue Ratio: max(Np,Nt) / min (Np,Nt)
Reproducibility Test: If Self-consistency Ratio >2 AND Rescue Ratio > 2, then 'Fail' else 'Pass'
Number of raw peaks
rep1
rep2
rep3
rep4
Number of peaks
5016
4433
5701
5866
Top 300000 raw peaks from spp with FDR 0.01
Peak calling statistics
Peak region size
rep1
rep2
rep3
rep4
idr_opt
overlap_opt
Min size
98.0
89.0
62.0
59.0
95.0
95.0
25 percentile
390.0
356.0
250.0
236.0
296.0
380.0
50 percentile (median)
390.0
356.0
250.0
236.0
373.0
380.0
75 percentile
390.0
356.0
250.0
236.0
392.0
380.0
Max size
1030.0
1149.0
794.0
803.0
2374.0
2374.0
Mean
374.25578149920256
350.53688247236636
244.4476407647781
231.83208319127175
426.7078774617068
390.9564183835182
Enrichment / Signal-to-noise ratio
Strand cross-correlation measures (trimmed/filtered SE BAM)
rep1
rep2
rep3
rep4
Number of Subsampled Reads
1923612
1905135
1188019
1793745
Estimated Fragment Length
145
150
95
105
Cross-correlation at Estimated Fragment Length
0.853646244568321
0.847652064872682
0.772256573959246
0.826749917164246
Phantom Peak
55
55
55
55
Cross-correlation at Phantom Peak
0.8490701
0.8365796
0.7626344
0.8148827
Argmin of Cross-correlation
1500
1500
1500
1500
Minimum of Cross-correlation
0.8227815
0.7990062
0.690895
0.763065
NSC (Normalized Strand Cross-correlation coeff.)
1.037513
1.060883
1.117763
1.083459
RSC (Relative Strand Cross-correlation coeff.)
1.174072
1.294688
1.134126
1.229019
Performed on subsampled (15000000) reads mapped from FASTQs that are trimmed to 50.
Such FASTQ trimming and subsampling reads are for cross-corrleation analysis only.
Untrimmed FASTQs are used for all the other analyses.
NOTE1: For SE datasets, reads from replicates are randomly subsampled to 15000000.
NOTE2: For PE datasets, the first end (R1) of each read-pair is selected and trimmed to 50 the reads are then randomly subsampled to 15000000.
Normalized strand cross-correlation coefficient (NSC) = col9 in outFile
Relative strand cross-correlation coefficient (RSC) = col10 in outFile
Estimated fragment length = col3 in outFile, take the top value
Peak enrichment
Fraction of reads in peaks (FRiP)
FRiP for spp raw peaks
rep1
rep2
rep3
rep4
rep1-pr1
rep2-pr1
rep3-pr1
rep4-pr1
rep1-pr2
rep2-pr2
rep3-pr2
rep4-pr2
pooled
pooled-pr1
pooled-pr2
Fraction of Reads in Peaks
0.2747165962990944
0.2909671039078806
0.4067404920176776
0.37124743158624257
0.33129113479027394
0.3401404523035794
0.4743794351008549
0.4180626785076323
0.33573141176850746
0.3374860567585971
0.4653973619788302
0.41816989176779273
0.2334488630943326
0.25579839869915794
0.2573024239569869
FRiP for overlap peaks
rep1_vs_rep2
rep1_vs_rep3
rep1_vs_rep4
rep2_vs_rep3
rep2_vs_rep4
rep3_vs_rep4
rep1-pr1_vs_rep1-pr2
rep2-pr1_vs_rep2-pr2
rep3-pr1_vs_rep3-pr2
rep4-pr1_vs_rep4-pr2
pooled-pr1_vs_pooled-pr2
Fraction of Reads in Peaks
0.15627457451443982
0.12677946341937152
0.1288988340024047
0.15248789698449491
0.1551004767294657
0.16487001451588368
0.2303016574778444
0.24178756873086119
0.30991892831013634
0.292205199787206
0.21130300897337723
FRiP for IDR peaks
rep1_vs_rep2
rep1_vs_rep3
rep1_vs_rep4
rep2_vs_rep3
rep2_vs_rep4
rep3_vs_rep4
rep1-pr1_vs_rep1-pr2
rep2-pr1_vs_rep2-pr2
rep3-pr1_vs_rep3-pr2
rep4-pr1_vs_rep4-pr2
pooled-pr1_vs_pooled-pr2
Fraction of Reads in Peaks
0.11355026371811168
0.09487889679284055
0.0943909516798977
0.11659385090322487
0.11855530001641523
0.1345695039246171
0.11990136691674998
0.1561645314643441
0.16556138172840024
0.1576349606867975
0.14606812066145677
For spp raw peaks:
repX: Peak from true replicate X
repX-prY: Peak from Yth pseudoreplicates from replicate X
pooled: Peak from pooled true replicates (pool of rep1, rep2, ...)
pooled-pr1: Peak from 1st pooled pseudo replicate (pool of rep1-pr1, rep2-pr1, ...)
pooled-pr2: Peak from 2nd pooled pseudo replicate (pool of rep1-pr2, rep2-pr2, ...)
For overlap/IDR peaks:
repX_vs_repY: Comparing two peaks from true replicates X and Y
repX-pr1_vs_repX-pr2: Comparing two peaks from both pseudoreplicates from replicate X
pooled-pr1_vs_pooled-pr2: Comparing two peaks from 1st and 2nd pooled pseudo replicates