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
31034981
27945511
68821547
Distinct Fragments
23550614
22852312
53412219
Positions with Two Read
3860164
3125220
9547729
NRF = Distinct/Total
0.758841
0.817745
0.776097
PBC1 = OneRead/Distinct
0.783605
0.83178
0.775564
PBC2 = OneRead/TwoRead
4.780726
6.082166
4.338686
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
25161
4650
N1
24963
5584
N2
21274
3370
Np
27554
6085
N optimal
27554
6085
N conservative
25161
4650
Optimal Set
pooled-pr1_vs_pooled-pr2
pooled-pr1_vs_pooled-pr2
Conservative Set
rep1_vs_rep2
rep1_vs_rep2
Rescue Ratio
1.0951075076507293
1.3086021505376344
Self Consistency Ratio
1.1734041553069474
1.656973293768546
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
34646
47338
Top 300000 raw peaks from spp with FDR 0.01
Peak calling statistics
Peak region size
rep1
rep2
idr_opt
overlap_opt
Min size
134.0
144.0
139.0
139.0
25 percentile
536.0
576.0
346.0
556.0
50 percentile (median)
536.0
576.0
556.0
556.0
75 percentile
536.0
576.0
556.0
556.0
Max size
6717.0
2966.0
6740.0
6740.0
Mean
532.4426773653524
570.4047277029025
563.4737880032868
554.2966538433622
Enrichment / Signal-to-noise ratio
Strand cross-correlation measures (trimmed/filtered SE BAM)
rep1
rep2
Number of Subsampled Reads
15000000
15000000
Estimated Fragment Length
190
225
Cross-correlation at Estimated Fragment Length
0.83728348962336
0.840592852464837
Phantom Peak
55
55
Cross-correlation at Phantom Peak
0.8243756
0.8339866
Argmin of Cross-correlation
1500
1500
Minimum of Cross-correlation
0.8017002
0.8232266
NSC (Normalized Strand Cross-correlation coeff.)
1.044385
1.021095
RSC (Relative Strand Cross-correlation coeff.)
1.569242
1.613958
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
0.35875874893367365
0.3856892819858446
Synthetic AUC
0.49815358590687836
0.49812555486794674
X-intercept
0.028393971612035152
0.02834391524856989
Synthetic X-intercept
0.0
0.0
Elbow Point
0.624525215392532
0.5624453134229142
Synthetic Elbow Point
0.4999625342001759
0.5025454152796675
JS Distance
0.1367693242129129
0.09215841265136616
Synthetic JS Distance
0.20891747639220346
0.17044952096665061
% Genome Enriched
24.589687988675248
27.015819813109562
Diff. Enrichment
14.148857841169239
9.492147171537113
CHANCE Divergence
0.12590952726974336
0.08398596253739275
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
0.27244705525347346
0.311337594872263
0.3060041503848416
0.2664975684921082
0.30391308050474697
0.26381495556799
0.25879017098343055
0.28573197504622444
0.2841050672995402
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
0.20102944774356887
0.22015506758830647
0.17199726409428143
0.21657689366418248
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
0.0653889708299371
0.08272432239121669
0.0465281537495392
0.0804544932339735
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