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
Total Fragments
3110116
3908713
455283
955496
9503613
Distinct Fragments
2961467
3699158
438921
909076
9014297
Positions with Two Read
125887
176881
15140
41305
407072
NRF = Distinct/Total
0.952205
0.946388
0.964062
0.951418
0.948513
PBC1 = OneRead/Distinct
0.954551
0.948796
0.964292
0.951944
0.951771
PBC2 = OneRead/TwoRead
22.455615
19.842414
27.955614
20.951192
21.076237
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
6976
52
N1
8853
32
N2
8445
20
N3
1285
31
N4
4353
42
Np
4412
54
N optimal
6976
54
N conservative
6976
52
Optimal Set
rep1_vs_rep2
pooled-pr1_vs_pooled-pr2
Conservative Set
rep1_vs_rep2
rep3_vs_rep4
Rescue Ratio
1.5811423390752493
1.0384615384615385
Self Consistency Ratio
6.889494163424125
2.1
Reproducibility Test
borderline
borderline
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
32626
28497
15179
22386
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
494.0
295.0
656.0
233.0
490.0
490.0
25 percentile
616.0
616.0
656.0
796.0
490.0
490.0
50 percentile (median)
616.0
616.0
656.0
796.0
490.0
490.0
75 percentile
616.0
616.0
656.0
796.0
10607.0
490.0
Max size
3853.0
1537.0
10679.0
4339.0
10607.0
10607.0
Mean
617.8282351498805
616.0776572972594
669.4046379866921
798.624050746002
4340.925925925926
519.8093463302753
Enrichment / Signal-to-noise ratio
Strand cross-correlation measures (trimmed/filtered SE BAM)
rep1
rep2
rep3
rep4
Number of Subsampled Reads
3110116
3908713
455283
955496
Estimated Fragment Length
135
110
150
100
Cross-correlation at Estimated Fragment Length
0.537100097510244
0.590448654945902
0.15326921867832
0.273077208765947
Phantom Peak
30
30
35
30
Cross-correlation at Phantom Peak
0.5370563
0.5905585
0.1536968
0.2733146
Argmin of Cross-correlation
1500
1500
1500
1500
Minimum of Cross-correlation
0.532113
0.5861845
0.1482078
0.2630626
NSC (Normalized Strand Cross-correlation coeff.)
1.009372
1.007274
1.034151
1.038069
RSC (Relative Strand Cross-correlation coeff.)
1.008865
0.9748779
0.9220954
0.9768445
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
Jensen-Shannon distance (filtered/deduped BAM)
rep1
rep2
rep3
rep4
AUC
0.37326712412900637
0.3815425597553023
0.2736959135377296
0.3229026792446403
Synthetic AUC
0.4935165088158732
0.4941953364289465
0.4833065546453137
0.48832965144624785
X-intercept
0.04006553856777075
0.03850291071866911
0.1619764548105189
0.06674197082695543
Synthetic X-intercept
9.377510238682775e-204
1.3058409724186177e-254
6.290012057667916e-30
4.608001870058969e-62
Elbow Point
0.483682213782657
0.4709636604208119
0.6179085533418882
0.5423735578621335
Synthetic Elbow Point
0.5002764899398032
0.49496913920425956
0.5068143687080133
0.5145862773703348
JS Distance
0.043375758712920105
0.0298563104931801
0.16577700727937048
0.11784325645776607
Synthetic JS Distance
0.1710576419100371
0.16288586592994708
0.2350742312565587
0.2150772580817568
% Genome Enriched
39.52650180909597
40.01923848335292
37.12727933120326
42.60465914556387
Diff. Enrichment
9.984993423150629
8.26258131010661
26.804787693090145
18.251039780689826
CHANCE Divergence
0.08490030513369343
0.07024092179558515
0.23080328764479074
0.15758235723288583
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
0.24291769323363904
0.2081820944440011
0.204134801243802
0.23514545403358014
0.2470731112284756
0.23524502641490777
0.20819529283752283
0.24329902265457673
0.24490827249890754
0.22958062851984007
0.139507521640474
0.21170692197911906
0.11974112234843502
0.14853278511840992
0.15231652539751284
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
0.05498867104019414
0.026673015322277383
0.03544155968055934
0.025746083555517718
0.03432914256893614
0.021482147932667457
0.07278214617956232
0.0658814469816842
0.04750952092391665
0.06468714334904853
0.03873810075628278
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
0.009352594276731513
0.009278598121809303
0.009423744425695177
0.009246178401759772
0.009367319264082253
0.009550948518103258
0.0073229188728361665
0.004858721449335409
0.029493853972691202
0.016428412100632145
0.009478560975244575
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