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Report generated at 2021-10-23 08:27:44

Pipeline type: Histone ChIP-Seq

Peak caller: MACS2

Alignment

Flagstat (raw BAM)

	rep1	ctl1
Total	164304580	186874232
Total(QC-failed)	0	0
Dupes	0	0
Dupes(QC-failed)	0	0
Mapped	161463393	183817074
Mapped(QC-failed)	0	0
% Mapped	98.2700	98.3600
Paired	164304580	186874232
Paired(QC-failed)	0	0
Read1	82152290	93437116
Read1(QC-failed)	0	0
Read2	82152290	93437116
Read2(QC-failed)	0	0
Properly Paired	159029347	181190078
Properly Paired(QC-failed)	0	0
% Properly Paired	96.7900	96.9600
With itself	160604010	182989360
With itself(QC-failed)	0	0
Singletons	859383	827714
Singletons(QC-failed)	0	0
% Singleton	0.5200	0.4400
Diff. Chroms	673235	1011019
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	66448228	79420006
Unmapped Reads	0	0
Unpaired Dupes	0	0
Paired Dupes	2750894	3323828
Paired Opt. Dupes	39140	42686
% Dupes/100	0.0414	0.0419

Library complexity (filtered non-mito BAM)

	rep1	ctl1
Total Read Pairs	66447920	79383916
Distinct Read Pairs	63697038	76061755
One Read Pair	61015379	72820637
Two Read Pairs	2614319	3164810
NRF = Distinct/Total	0.9586	0.9582
PBC1 = OnePair/Distinct	0.9579	0.9574
PBC2 = OnePair/TwoPair	23.3389	23.0095

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	127394668	152192356
Total(QC-failed)	0	0
Dupes	0	0
Dupes(QC-failed)	0	0
Mapped	127394668	152192356
Mapped(QC-failed)	0	0
% Mapped	100.0000	100.0000
Paired	127394668	152192356
Paired(QC-failed)	0	0
Read1	63697334	76096178
Read1(QC-failed)	0	0
Read2	63697334	76096178
Read2(QC-failed)	0	0
Properly Paired	127394668	152192356
Properly Paired(QC-failed)	0	0
% Properly Paired	100.0000	100.0000
With itself	127394668	152192356
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	231031
Np	0
N optimal	231031
N conservative	231031
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.	130
Corr. Est. Fragment Len.	0.1842
Phantom Peak	50
Corr. Phantom Peak	0.1973
Argmin. Corr.	1500
Min. Corr.	0.1796
NSC	1.0255
RSC	0.2584

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	0.2695

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	0.2151
AUC	0.4964
CHANCE divergence	0.1500
Elbow Point	0.0000
JS Distance	0.5930
Synthetic AUC	0.5057
Synthetic Elbow Point	0.2698
Synthetic JS Distance	0.3886