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
Total Fragments
16864996
14426546
56089165
Distinct Fragments
14739982
12798847
43899072
Positions with Two Read
1562289
1219494
7518324
NRF = Distinct/Total
0.873999
0.887173
0.782666
PBC1 = OneRead/Distinct
0.878134
0.89188
0.784732
PBC2 = OneRead/TwoRead
8.285074
9.360465
4.582009
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
6676
211
N1
6534
136
N2
7005
124
Np
6331
223
N optimal
6676
223
N conservative
6676
211
Optimal Set
rep1_vs_rep2
pooled-pr1_vs_pooled-pr2
Conservative Set
rep1_vs_rep2
rep1_vs_rep2
Rescue Ratio
1.054493760859264
1.0568720379146919
Self Consistency Ratio
1.0720844811753902
1.096774193548387
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
24178
16963
Top 300000 raw peaks from spp with FDR 0.01
Peak calling statistics
Peak region size
rep1
rep2
idr_opt
overlap_opt
Min size
188.0
251.0
195.0
195.0
25 percentile
750.0
816.0
776.0
776.0
50 percentile (median)
750.0
816.0
776.0
776.0
75 percentile
750.0
816.0
776.0
776.0
Max size
3588.0
2792.0
3622.0
3622.0
Mean
751.7483662833981
816.5427106054353
912.4215246636771
780.2633313361295
Enrichment / Signal-to-noise ratio
Strand cross-correlation measures (trimmed/filtered SE BAM)
rep1
rep2
Number of Subsampled Reads
15000000
14426546
Estimated Fragment Length
285
300
Cross-correlation at Estimated Fragment Length
0.834005316973396
0.828932885097844
Phantom Peak
50
50
Cross-correlation at Phantom Peak
0.8348181
0.8296212
Argmin of Cross-correlation
1500
1500
Minimum of Cross-correlation
0.8313625
0.8257135
NSC (Normalized Strand Cross-correlation coeff.)
1.003179
1.003899
RSC (Relative Strand Cross-correlation coeff.)
0.7647855
0.8238497
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
0.42224759265341405
0.41047754908288714
Synthetic AUC
0.4976670929508647
0.49749647797563695
X-intercept
0.02853467711444478
0.02865470520101704
Synthetic X-intercept
0.0
0.0
Elbow Point
0.3853981831748629
0.4220727650270163
Synthetic Elbow Point
0.5014102831823137
0.49807101578266066
JS Distance
0.014144205111149633
0.02624001480190002
Synthetic JS Distance
0.12308485417386447
0.13465314952213486
% Genome Enriched
36.75280015523632
38.82768567844876
Diff. Enrichment
3.238964516747167
4.17860243810585
CHANCE Divergence
0.027696841811268774
0.03552757502904798
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
0.1855889778486601
0.15274347219304535
0.18336271729824913
0.19937533575079106
0.16570902813908225
0.20057974660706365
0.13898482542789897
0.14608360946445542
0.15433812927774748
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
0.05560334550435612
0.05090678748583261
0.06379210404661446
0.052324988867347745
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
0.0037746926724262684
0.0029248214566685565
0.0030038478094143507
0.004004236754863966
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