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
ctl1
ctl2
Total Fragments
9740837
8643393
9369623
7434022
Distinct Fragments
8017538
7371588
7900285
6534148
Positions with Two Read
1247448
989329
1103313
742304
NRF = Distinct/Total
0.823085
0.852858
0.843181
0.878952
PBC1 = OneRead/Distinct
0.817007
0.847763
0.838709
0.874892
PBC2 = OneRead/TwoRead
5.251028
6.316763
6.005589
7.701261
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
31497
3165
N1
23552
2219
N2
21667
2733
Np
31009
3250
N optimal
31497
3250
N conservative
31497
3165
Optimal Set
rep1_vs_rep2
pooled-pr1_vs_pooled-pr2
Conservative Set
rep1_vs_rep2
rep1_vs_rep2
Rescue Ratio
1.0157373665709954
1.0268562401263823
Self Consistency Ratio
1.086998661559053
1.231635872014421
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
Number of peaks
67615
58604
Top 300000 raw peaks from spp with FDR 0.01
Peak calling statistics
Peak region size
rep1
rep2
idr_opt
overlap_opt
Min size
78.0
85.0
85.0
85.0
25 percentile
310.0
340.0
238.0
340.0
50 percentile (median)
310.0
340.0
340.0
340.0
75 percentile
310.0
340.0
340.0
340.0
Max size
771.0
909.0
933.0
933.0
Mean
308.52100865192637
338.01527199508564
300.9913846153846
335.7991237260691
Enrichment / Signal-to-noise ratio
Strand cross-correlation measures (trimmed/filtered SE BAM)
rep1
rep2
Number of Subsampled Reads
9170147
8107005
Estimated Fragment Length
155
165
Cross-correlation at Estimated Fragment Length
0.724277268368307
0.713007108430159
Phantom Peak
55
55
Cross-correlation at Phantom Peak
0.7149897
0.6959266
Argmin of Cross-correlation
1500
1500
Minimum of Cross-correlation
0.7021627
0.6757287
NSC (Normalized Strand Cross-correlation coeff.)
1.031495
1.055168
RSC (Relative Strand Cross-correlation coeff.)
1.724062
1.845657
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
AUC
0.3955609215900333
0.3833856379536129
Synthetic AUC
0.4984093244193228
0.49834402557357244
X-intercept
0.01942348385036132
0.01939154389747275
Synthetic X-intercept
0.0
0.0
Elbow Point
0.598486844731904
0.638248093584062
Synthetic Elbow Point
0.5002320826023485
0.5019624596770786
JS Distance
0.10448174059081353
0.13786657671355673
Synthetic JS Distance
0.17308752927514054
0.19933580619647062
% Genome Enriched
34.442248572683354
30.09082924102687
Diff. Enrichment
10.508067278762745
12.246396003334503
CHANCE Divergence
0.09031797077714677
0.10679047467004008
Peak enrichment
Fraction of reads in peaks (FRiP)
FRiP for spp raw peaks
rep1
rep2
rep1-pr1
rep2-pr1
rep1-pr2
rep2-pr2
pooled
pooled-pr1
pooled-pr2
Fraction of Reads in Peaks
0.3674289076002125
0.3665189490269419
0.3475704985652575
0.35750839821738883
0.3418544600822379
0.3631858093773563
0.4429533504045434
0.37923274680036156
0.3501717474366653
FRiP for overlap peaks
rep1_vs_rep2
rep1-pr1_vs_rep1-pr2
rep2-pr1_vs_rep2-pr2
pooled-pr1_vs_pooled-pr2
Fraction of Reads in Peaks
0.22053917251285404
0.16926375164535992
0.19107660687694417
0.21850003581576372
FRiP for IDR peaks
rep1_vs_rep2
rep1-pr1_vs_rep1-pr2
rep2-pr1_vs_rep2-pr2
pooled-pr1_vs_pooled-pr2
Fraction of Reads in Peaks
0.0679763459366644
0.04846183292691603
0.07644729054453961
0.06877335182940904
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