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
32736011
20763364
34473281
31624735
Distinct Fragments
28413454
18267691
29624682
26276588
Positions with Two Read
3368828
2001379
3715227
3845088
NRF = Distinct/Total
0.867957
0.879804
0.859352
0.830887
PBC1 = OneRead/Distinct
0.865647
0.877616
0.856681
0.827295
PBC2 = OneRead/TwoRead
7.301065
8.010485
6.831048
5.653576
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
19538
84
N1
13785
22
N2
11476
27
N3
14039
34
Np
25042
200
N optimal
25042
200
N conservative
19538
84
Optimal Set
pooled-pr1_vs_pooled-pr2
pooled-pr1_vs_pooled-pr2
Conservative Set
rep1_vs_rep3
rep1_vs_rep3
Rescue Ratio
1.2817074419080765
2.380952380952381
Self Consistency Ratio
1.2233356570233531
1.5454545454545454
Reproducibility Test
pass
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
298411
298413
298374
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
170.0
164.0
164.0
170.0
170.0
25 percentile
170.0
164.0
164.0
170.0
170.0
50 percentile (median)
170.0
164.0
164.0
170.0
170.0
75 percentile
170.0
164.0
164.0
170.0
170.0
Max size
170.0
164.0
164.0
170.0
170.0
Mean
170.0
164.0
164.0
170.0
170.0
Enrichment / Signal-to-noise ratio
Strand cross-correlation measures (trimmed/filtered SE BAM)
rep1
rep2
rep3
Number of Subsampled Reads
15000000
15000000
15000000
Estimated Fragment Length
105
100
95
Cross-correlation at Estimated Fragment Length
0.153652446835153
0.147559845342375
0.153657659084962
Phantom Peak
50
50
50
Cross-correlation at Phantom Peak
0.1551925
0.1488867
0.155069
Argmin of Cross-correlation
1500
1500
1500
Minimum of Cross-correlation
0.1529773
0.1466717
0.1528687
NSC (Normalized Strand Cross-correlation coeff.)
1.004413
1.006056
1.005161
RSC (Relative Strand Cross-correlation coeff.)
0.304764
0.4009805
0.358584
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
0.3218989114258676
0.29846365966314364
0.3238289631588766
Synthetic AUC
0.4934532344548082
0.49183471656775957
0.49358712248955566
X-intercept
0.10328364415725665
0.12711925402258406
0.10228799462589169
Synthetic X-intercept
8.357599947009978e-200
6.031549057200959e-128
2.581348473034729e-208
Elbow Point
0.5179136943795783
0.5209226352323982
0.512753510764211
Synthetic Elbow Point
0.5024001770515042
0.5006989617661322
0.5089331818356684
JS Distance
0.01054439451219131
0.028048750961032733
0.01256720154184957
Synthetic JS Distance
0.2042938761614347
0.22451334818584007
0.202140348089327
% Genome Enriched
38.39388535235597
38.72536867022808
38.9908752119254
Diff. Enrichment
16.240461198660118
20.661031795802664
16.284010539699572
CHANCE Divergence
0.13885050855931355
0.17775386137648294
0.1393181680354296
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
0.0658057549579386
0.07770794765419757
0.06386988250215446
0.08643548777841856
0.10447900721818104
0.08413027409964578
0.0864465188475301
0.1045711363424025
0.08424809606509676
0.0489682274776191
0.05860485587987521
0.058688613582286546
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
0.003184362944476541
0.0040454176880077115
0.003219268978309873
0.0040466633896662635
0.004021380267393759
0.004041610122567862
0.005275494024370861
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
3.119109525035815e-05
4.147258020182869e-05
3.310742800530781e-05
1.4421190669710961e-05
1.9614623461242352e-05
2.2186378109655897e-05
9.517568003422348e-05
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