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Report generated at 2021-12-31 12:03:34

Pipeline type: Histone ChIP-Seq

Peak caller: MACS2

Alignment

Flagstat (raw BAM)

	rep1	ctl1
Total	72656920	73857574
Total(QC-failed)	0	0
Dupes	0	0
Dupes(QC-failed)	0	0
Mapped	71346631	71880916
Mapped(QC-failed)	0	0
% Mapped	98.2000	97.3200
Paired	72656920	73857574
Paired(QC-failed)	0	0
Read1	36328460	36928787
Read1(QC-failed)	0	0
Read2	36328460	36928787
Read2(QC-failed)	0	0
Properly Paired	68601813	56693280
Properly Paired(QC-failed)	0	0
% Properly Paired	94.4200	76.7600
With itself	70857133	70763351
With itself(QC-failed)	0	0
Singletons	489498	1117565
Singletons(QC-failed)	0	0
% Singleton	0.6700	1.5100
Diff. Chroms	1891202	11478644
Diff. Chroms (QC-failed)	0	0

Marking duplicates (filtered BAM)

Filtered out (samtools view -F 1804):

read unmapped (0x4)
mate unmapped (0x8)
not primary alignment (0x100)
read fails platform/vendor quality checks (0x200)
read is PCR or optical duplicate (0x400)

	rep1	ctl1
Unpaired Reads	0	0
Paired Reads	31097205	24996827
Unmapped Reads	0	0
Unpaired Dupes	0	0
Paired Dupes	1668167	241052
Paired Opt. Dupes	1230	1015
% Dupes/100	0.0536	0.0096

Library complexity (filtered non-mito BAM)

	rep1	ctl1
Total Read Pairs	31090410	24993555
Distinct Read Pairs	29422886	24752593
One Read Pair	27853006	24516128
Two Read Pairs	1482125	233547
NRF = Distinct/Total	0.9464	0.9904
PBC1 = OnePair/Distinct	0.9466	0.9904
PBC2 = OnePair/TwoPair	18.7926	104.9730

Mitochondrial reads are filtered out.

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

Flagstat (filtered/deduped BAM)

Filtered and duplicates removed

	rep1	ctl1
Total	58858076	49511550
Total(QC-failed)	0	0
Dupes	0	0
Dupes(QC-failed)	0	0
Mapped	58858076	49511550
Mapped(QC-failed)	0	0
% Mapped	100.0000	100.0000
Paired	58858076	49511550
Paired(QC-failed)	0	0
Read1	29429038	24755775
Read1(QC-failed)	0	0
Read2	29429038	24755775
Read2(QC-failed)	0	0
Properly Paired	58858076	49511550
Properly Paired(QC-failed)	0	0
% Properly Paired	100.0000	100.0000
With itself	58858076	49511550
With itself(QC-failed)	0	0
Singletons	0	0
Singletons(QC-failed)	0	0
% Singleton	0.0000	0.0000
Diff. Chroms	0	0
Diff. Chroms (QC-failed)	0	0

Peak calling

Reproducibility QC and peak detection statistics

The number of peaks is capped at 300K for peak-caller MACS2

	overlap
Nt	0
N1	224359
Np	0
N optimal	224359
N conservative	224359
Optimal Set	rep1-pr
Conservative Set	rep1-pr
Rescue Ratio	0.0000
Self Consistency Ratio	1.0000
Reproducibility	pass

Overlapping peaks

N1: Replicate 1 self-consistent overlapping peaks (comparing two pseudoreplicates generated by subsampling Rep1 reads)
N2: Replicate 2 self-consistent overlapping peaks (comparing two pseudoreplicates generated by subsampling Rep2 reads)
Nt: True Replicate consisten overlapping peaks (comparing true replicates Rep1 vs Rep2 )
Np: Pooled-pseudoreplicate consistent overlapping 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'

Enrichment

Strand cross-correlation measures

Performed on subsampled reads (15M)

	rep1
Reads	15000000
Est. Fragment Len.	150
Corr. Est. Fragment Len.	0.2054
Phantom Peak	55
Corr. Phantom Peak	0.1808
Argmin. Corr.	1500
Min. Corr.	0.1726
NSC	1.1900
RSC	4.0022

NOTE1: For SE datasets, reads from replicates are randomly subsampled.
NOTE2: For PE datasets, the first end of each read-pair is selected and the reads are then randomly subsampled.

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

Fraction of reads in overlapping peaks

	rep1-pr
Fraction of Reads in Peak	0.3811

ppr: Overlapping peaks comparing pooled pseudo replicates
rep1-pr: Overlapping peaks comparing pseudoreplicates from replicate 1
rep2-pr: Overlapping peaks comparing pseudoreplicates from replicate 2
repi-repj: Overlapping peaks comparing true replicates (rep i vs. rep j)

Other quality metrics

Fingerprint and Jensen-Shannon distance

	rep1
% genome enriched	0.1398
AUC	0.4947
CHANCE divergence	0.3639
Elbow Point	0.0000
JS Distance	0.6862
Synthetic AUC	0.5030
Synthetic Elbow Point	0.2475
Synthetic JS Distance	0.4272