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
ctl1
Total Fragments
999706
1768510
443040
6590671
Distinct Fragments
950557
1709853
415914
6407757
Positions with Two Read
43533
50584
24171
142740
NRF = Distinct/Total
0.950837
0.966833
0.938773
0.972247
PBC1 = OneRead/Distinct
0.951515
0.96851
0.938499
0.976353
PBC2 = OneRead/TwoRead
20.776629
32.737822
16.148897
43.82955
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
6345
203
N1
3532
37
N2
7459
85
N3
1242
105
Np
6623
385
N optimal
6623
385
N conservative
6345
203
Optimal Set
pooled-pr1_vs_pooled-pr2
pooled-pr1_vs_pooled-pr2
Conservative Set
rep1_vs_rep2
rep1_vs_rep3
Rescue Ratio
1.0438140267927503
1.896551724137931
Self Consistency Ratio
6.005636070853462
2.8378378378378377
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
Number of peaks
30075
27993
13618
Top 300000 raw peaks from spp with FDR 0.01
Peak calling statistics
Peak region size
rep1
rep2
rep3
idr_opt
overlap_opt
Min size
624.0
223.0
155.0
120.0
120.0
25 percentile
624.0
630.0
430.0
480.0
480.0
50 percentile (median)
624.0
630.0
430.0
480.0
480.0
75 percentile
624.0
630.0
430.0
480.0
480.0
Max size
624.0
630.0
430.0
480.0
480.0
Mean
624.0
629.9711356410531
429.92443824350124
453.53246753246754
478.4614223161709
Enrichment / Signal-to-noise ratio
Strand cross-correlation measures (trimmed/filtered SE BAM)
rep1
rep2
rep3
Number of Subsampled Reads
999706
1768510
443040
Estimated Fragment Length
105
110
115
Cross-correlation at Estimated Fragment Length
0.285162344153938
0.409146148372385
0.161920680611905
Phantom Peak
30
30
30
Cross-correlation at Phantom Peak
0.2838761
0.4084271
0.1568671
Argmin of Cross-correlation
1500
1500
1500
Minimum of Cross-correlation
0.2723465
0.400421
0.1432267
NSC (Normalized Strand Cross-correlation coeff.)
1.047057
1.02179
1.13052
RSC (Relative Strand Cross-correlation coeff.)
1.111556
1.089813
1.370483
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
AUC
0.32500957532560815
0.35600126577304697
0.24411290330190194
Synthetic AUC
0.48851279362384387
0.49143730588890844
0.48264944382571995
X-intercept
0.06554806272694626
0.04681576952236543
0.20679741430908583
Synthetic X-intercept
4.311188917956582e-64
3.106909242160553e-116
1.1741059825216368e-27
Elbow Point
0.5268624556083157
0.5179482063764415
0.6752424085232034
Synthetic Elbow Point
0.4915263523191996
0.5022414555862252
0.5220221409584308
JS Distance
0.11060020150808281
0.07032658538160727
0.19146154263851656
Synthetic JS Distance
0.20579162137267534
0.17821394160646323
0.255485241322804
% Genome Enriched
37.038027213598816
39.69294920394238
31.443278400702287
Diff. Enrichment
17.636888929010873
12.895472225206827
30.68840829478092
CHANCE Divergence
0.15033593436312406
0.10983569407538314
0.26302709023136334
Peak enrichment
Fraction of reads in peaks (FRiP)
FRiP for spp raw peaks
rep1
rep2
rep3
rep1-pr1
rep2-pr1
rep3-pr1
rep1-pr2
rep2-pr2
rep3-pr2
pooled
pooled-pr1
pooled-pr2
Fraction of Reads in Peaks
0.26661020346692443
0.23015234882046592
0.21678532036498546
0.19521973397254927
0.2583616336483349
0.13611512989834457
0.1917462889575497
0.26383320623479634
0.14201200383893656
0.17386380363403864
0.21998039091082403
0.22276900929876664
FRiP for overlap peaks
rep1_vs_rep2
rep1_vs_rep3
rep2_vs_rep3
rep1-pr1_vs_rep1-pr2
rep2-pr1_vs_rep2-pr2
rep3-pr1_vs_rep3-pr2
pooled-pr1_vs_pooled-pr2
Fraction of Reads in Peaks
0.06166070417685542
0.047494035889065915
0.048307515330958714
0.04434250283103125
0.06806035094357471
0.05258722359895528
0.06879007820945036
FRiP for IDR peaks
rep1_vs_rep2
rep1_vs_rep3
rep2_vs_rep3
rep1-pr1_vs_rep1-pr2
rep2-pr1_vs_rep2-pr2
rep3-pr1_vs_rep3-pr2
pooled-pr1_vs_pooled-pr2
Fraction of Reads in Peaks
0.005703758605730604
0.006652763917177482
0.006240674490853949
0.0018250837240103984
0.002824972570672487
0.012175731618226354
0.010993806662296119
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