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
Total Fragments
10986820
8733387
11000413
15244528
26050027
15632616
Distinct Fragments
10111032
8139353
10263303
13982738
25756642
15536775
Positions with Two Read
420952
279686
346218
589548
250069
74630
NRF = Distinct/Total
0.920287
0.931981
0.932993
0.91723
0.988738
0.993869
PBC1 = OneRead/Distinct
0.943274
0.952131
0.953074
0.942231
0.990055
0.99506
PBC2 = OneRead/TwoRead
22.656904
27.708673
28.252979
22.347575
101.973839
207.155608
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
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
29683
33292
25762
27043
Top 500000 raw peaks from macs2 with p-val threshold 0.01
Peak calling statistics
Peak region size
rep1
rep2
rep3
rep4
idr_opt
overlap_opt
Min size
210.0
215.0
260.0
255.0
237.0
235.0
25 percentile
233.0
215.0
291.0
298.0
682.0
433.0
50 percentile (median)
329.0
291.0
407.0
420.0
940.0
577.0
75 percentile
521.0
474.0
624.0
626.0
1347.0
859.0
Max size
4909.0
4582.0
4614.0
4573.0
4873.0
4873.0
Mean
449.2894586126739
417.3429352396972
527.1118701964133
532.3520319491181
1052.2219728145653
719.7464756288584
rep1rep2rep3rep4idr_optoverlap_opt
Enrichment / Signal-to-noise ratio
Strand cross-correlation measures (trimmed/filtered SE BAM)
rep1
rep2
rep3
rep4
Number of Subsampled Reads
10986820
8733387
11000413
15000000
Estimated Fragment Length
210
215
260
255
Cross-correlation at Estimated Fragment Length
0.311374246136933
0.307192162984864
0.312487328340206
0.321756914810112
Phantom Peak
55
55
55
55
Cross-correlation at Phantom Peak
0.2281194
0.2161116
0.2202469
0.2407842
Argmin of Cross-correlation
1500
1500
1500
1500
Minimum of Cross-correlation
0.09853433
0.08147826
0.1003748
0.1291353
NSC (Normalized Strand Cross-correlation coeff.)
3.160058
3.770234
3.113206
2.491626
RSC (Relative Strand Cross-correlation coeff.)
1.642472
1.676508
1.76949
1.725244
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
rep1rep2rep3rep4
Jensen-Shannon distance (filtered/deduped BAM)
rep1
rep2
rep3
rep4
AUC
0.14262390146148074
0.12991214774798435
0.14699865106462767
0.16272972025202972
Synthetic AUC
0.4851472148595775
0.4833392185062684
0.48516243210108906
0.4874118172204721
X-intercept
0.36852375373370067
0.4343308528632036
0.35965484011563975
0.27658574815964043
Synthetic X-intercept
5.431627859921942e-38
4.7645129304390755e-30
4.515915389430275e-38
3.256672764389922e-53
Elbow Point
0.6840799401808171
0.7372214469364661
0.6773642772315235
0.7226825064877422
Synthetic Elbow Point
0.5145378611638363
0.4935369347245437
0.5067026686995642
0.49888640971542897
JS Distance
0.27866655512848626
0.2911584257810322
0.27136141102099504
0.26142242560270856
Synthetic JS Distance
0.4350023063292984
0.4316912421807552
0.4287946906802517
0.4348209789360988
% Genome Enriched
31.58940688640353
25.88119334471604
32.482496091360204
27.11836280974221
Diff. Enrichment
27.003109435389955
30.33820897059355
26.497411934071508
22.583472823541516
CHANCE Divergence
0.2527705841409282
0.27389923843950725
0.2488107962135242
0.20098939583964912
Peak enrichment
Fraction of reads in peaks (FRiP)
FRiP for macs2 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
0.23498313144129318
0.24771845411459326
0.22631338895088296
0.215965061993726
0.22812087858844
0.25494423813421957
0.21978939724037763
0.21118523903328631
0.2278818134237486
0.25532868254335755
0.21950156756469524
0.2109619060911155
0.24363551984402376
0.2363682646562558
0.23619825969098143
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
0.22309486922975347
0.22304579383249662
0.22477674563042943
0.22203127923788685
0.22326609254882201
0.22333693919142783
0.21591293697500394
0.22601743934305513
0.20866393892697097
0.1998543164358361
0.23213180155062568
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
0.19850970631861956
0.19852251043954533
0.20019213389006385
0.19705052410722365
0.1991374898237447
0.19960522181729778
0.19087188148189166
0.20310515732785953
0.18594177869883274
0.14117974700936248
0.18496173804324584
For macs2 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