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Report generated at 2022-09-02 16:29:47

Pipeline type: Histone ChIP-Seq

Peak caller: MACS2

Alignment

Flagstat (raw BAM)

	rep1	ctl1
Total	83211848	126123212
Total(QC-failed)	0	0
Dupes	0	0
Dupes(QC-failed)	0	0
Mapped	81481009	123585796
Mapped(QC-failed)	0	0
% Mapped	97.9200	97.9900
Paired	83211848	126123212
Paired(QC-failed)	0	0
Read1	41605924	63061606
Read1(QC-failed)	0	0
Read2	41605924	63061606
Read2(QC-failed)	0	0
Properly Paired	80588826	122490924
Properly Paired(QC-failed)	0	0
% Properly Paired	96.8500	97.1200
With itself	81122383	122901660
With itself(QC-failed)	0	0
Singletons	358626	684136
Singletons(QC-failed)	0	0
% Singleton	0.4300	0.5400
Diff. Chroms	89453	61311
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	35698817	52026374
Unmapped Reads	0	0
Unpaired Dupes	0	0
Paired Dupes	1681302	1378590
Paired Opt. Dupes	2922	7099
% Dupes/100	0.0471	0.0265

Library complexity (filtered non-mito BAM)

	rep1	ctl1
Total Read Pairs	35696127	51966555
Distinct Read Pairs	34014943	50590836
One Read Pair	32400547	49267292
Two Read Pairs	1550241	1284538
NRF = Distinct/Total	0.9529	0.9735
PBC1 = OnePair/Distinct	0.9525	0.9738
PBC2 = OnePair/TwoPair	20.9003	38.3541

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	68035030	101295568
Total(QC-failed)	0	0
Dupes	0	0
Dupes(QC-failed)	0	0
Mapped	68035030	101295568
Mapped(QC-failed)	0	0
% Mapped	100.0000	100.0000
Paired	68035030	101295568
Paired(QC-failed)	0	0
Read1	34017515	50647784
Read1(QC-failed)	0	0
Read2	34017515	50647784
Read2(QC-failed)	0	0
Properly Paired	68035030	101295568
Properly Paired(QC-failed)	0	0
% Properly Paired	100.0000	100.0000
With itself	68035030	101295568
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	50508
Np	0
N optimal	50508
N conservative	50508
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.	155
Corr. Est. Fragment Len.	0.1769
Phantom Peak	50
Corr. Phantom Peak	0.1855
Argmin. Corr.	1500
Min. Corr.	0.1706
NSC	1.0367
RSC	0.4195

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	0.0810

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	0.3032
AUC	0.4951
CHANCE divergence	0.1022
Elbow Point	0.0000
JS Distance	0.5624
Synthetic AUC	0.5078
Synthetic Elbow Point	0.1027
Synthetic JS Distance	0.2462