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Report generated at 2021-10-23 14:04:57

Pipeline type: Histone ChIP-Seq

Peak caller: MACS2

Alignment

Flagstat (raw BAM)

	rep1	ctl1
Total	161482978	186874232
Total(QC-failed)	0	0
Dupes	0	0
Dupes(QC-failed)	0	0
Mapped	159626659	183817074
Mapped(QC-failed)	0	0
% Mapped	98.8500	98.3600
Paired	161482978	186874232
Paired(QC-failed)	0	0
Read1	80741489	93437116
Read1(QC-failed)	0	0
Read2	80741489	93437116
Read2(QC-failed)	0	0
Properly Paired	158677718	181190078
Properly Paired(QC-failed)	0	0
% Properly Paired	98.2600	96.9600
With itself	159086594	182989360
With itself(QC-failed)	0	0
Singletons	540065	827714
Singletons(QC-failed)	0	0
% Singleton	0.3300	0.4400
Diff. Chroms	195457	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	73083007	79420006
Unmapped Reads	0	0
Unpaired Dupes	0	0
Paired Dupes	8292782	3323828
Paired Opt. Dupes	37710	42686
% Dupes/100	0.1135	0.0419

Library complexity (filtered non-mito BAM)

	rep1	ctl1
Total Read Pairs	73082315	79383916
Distinct Read Pairs	64789608	76061755
One Read Pair	57276288	72820637
Two Read Pairs	6798621	3164810
NRF = Distinct/Total	0.8865	0.9582
PBC1 = OnePair/Distinct	0.8840	0.9574
PBC2 = OnePair/TwoPair	8.4247	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	129580450	152192356
Total(QC-failed)	0	0
Dupes	0	0
Dupes(QC-failed)	0	0
Mapped	129580450	152192356
Mapped(QC-failed)	0	0
% Mapped	100.0000	100.0000
Paired	129580450	152192356
Paired(QC-failed)	0	0
Read1	64790225	76096178
Read1(QC-failed)	0	0
Read2	64790225	76096178
Read2(QC-failed)	0	0
Properly Paired	129580450	152192356
Properly Paired(QC-failed)	0	0
% Properly Paired	100.0000	100.0000
With itself	129580450	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	123537
Np	0
N optimal	123537
N conservative	123537
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.	235
Corr. Est. Fragment Len.	0.2208
Phantom Peak	50
Corr. Phantom Peak	0.2200
Argmin. Corr.	1500
Min. Corr.	0.1966
NSC	1.1231
RSC	1.0376

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	0.7150

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	0.1071
AUC	0.4964
CHANCE divergence	0.1324
Elbow Point	0.0000
JS Distance	0.8510
Synthetic AUC	0.4987
Synthetic Elbow Point	0.5243
Synthetic JS Distance	0.6006