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Report generated at 2021-12-31 14:58:16

Pipeline type: Histone ChIP-Seq

Peak caller: MACS2

Alignment

Flagstat (raw BAM)

	rep1	ctl1
Total	125427566	265917970
Total(QC-failed)	0	0
Dupes	0	0
Dupes(QC-failed)	0	0
Mapped	117228809	247468204
Mapped(QC-failed)	0	0
% Mapped	93.4600	93.0600
Paired	125427566	265917970
Paired(QC-failed)	0	0
Read1	62713783	132958985
Read1(QC-failed)	0	0
Read2	62713783	132958985
Read2(QC-failed)	0	0
Properly Paired	110241734	230014570
Properly Paired(QC-failed)	0	0
% Properly Paired	87.8900	86.5000
With itself	111773547	235762704
With itself(QC-failed)	0	0
Singletons	5455262	11705500
Singletons(QC-failed)	0	0
% Singleton	4.3500	4.4000
Diff. Chroms	292655	1000726
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	45038879	94716882
Unmapped Reads	0	0
Unpaired Dupes	0	0
Paired Dupes	13161637	28466613
Paired Opt. Dupes	3176	5225
% Dupes/100	0.2922	0.3005

Library complexity (filtered non-mito BAM)

	rep1	ctl1
Total Read Pairs	45035316	94358262
Distinct Read Pairs	31874660	66006294
One Read Pair	22049069	44764704
Two Read Pairs	7233620	15675485
NRF = Distinct/Total	0.7078	0.6995
PBC1 = OnePair/Distinct	0.6917	0.6782
PBC2 = OnePair/TwoPair	3.0481	2.8557

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	63754484	132500538
Total(QC-failed)	0	0
Dupes	0	0
Dupes(QC-failed)	0	0
Mapped	63754484	132500538
Mapped(QC-failed)	0	0
% Mapped	100.0000	100.0000
Paired	63754484	132500538
Paired(QC-failed)	0	0
Read1	31877242	66250269
Read1(QC-failed)	0	0
Read2	31877242	66250269
Read2(QC-failed)	0	0
Properly Paired	63754484	132500538
Properly Paired(QC-failed)	0	0
% Properly Paired	100.0000	100.0000
With itself	63754484	132500538
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	119647
Np	0
N optimal	119647
N conservative	119647
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.	0
Corr. Est. Fragment Len.	0.1575
Phantom Peak	50
Corr. Phantom Peak	0.1631
Argmin. Corr.	1500
Min. Corr.	0.1545
NSC	1.0195
RSC	0.3481

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	0.2700

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	0.2228
AUC	0.4937
CHANCE divergence	0.1275
Elbow Point	0.0000
JS Distance	0.7074
Synthetic AUC	0.5026
Synthetic Elbow Point	0.2153
Synthetic JS Distance	0.3606