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Report generated at 2022-10-28 15:08:32

Pipeline type: Histone ChIP-Seq

Peak caller: MACS2

Alignment

Flagstat (raw BAM)

	rep1	ctl1
Total	60922837	132967577
Total(QC-failed)	0	0
Dupes	0	0
Dupes(QC-failed)	0	0
Mapped	46333960	120327666
Mapped(QC-failed)	0	0
% Mapped	76.0500	90.4900
Paired	0	0
Paired(QC-failed)	0	0
Read1	0	0
Read1(QC-failed)	0	0
Read2	0	0
Read2(QC-failed)	0	0
Properly Paired	0	0
Properly Paired(QC-failed)	0	0
% Properly Paired	0.0000	0.0000
With itself	0	0
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

Marking duplicates (filtered BAM)

Filtered out (samtools view -F 1804):

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

	rep1	ctl1
Unpaired Reads	40001820	101662952
Paired Reads	0	0
Unmapped Reads	0	0
Unpaired Dupes	2137889	2166997
Paired Dupes	0	0
Paired Opt. Dupes	0	0
% Dupes/100	0.0534	0.0213

Library complexity (filtered non-mito BAM)

	rep1	ctl1
Total Reads	39999997	101616265
Distinct Reads	38505466	99902404
One Read	37109231	98381677
Two Reads	1317192	1482009
NRF = Distinct/Total	0.9626	0.9831
PBC1 = OneRead/Distinct	0.9637	0.9848
PBC2 = OneRead/TwoReads	28.1730	66.3840

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	37863931	99495955
Total(QC-failed)	0	0
Dupes	0	0
Dupes(QC-failed)	0	0
Mapped	37863931	99495955
Mapped(QC-failed)	0	0
% Mapped	100.0000	100.0000
Paired	0	0
Paired(QC-failed)	0	0
Read1	0	0
Read1(QC-failed)	0	0
Read2	0	0
Read2(QC-failed)	0	0
Properly Paired	0	0
Properly Paired(QC-failed)	0	0
% Properly Paired	0.0000	0.0000
With itself	0	0
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	154489
Np	0
N optimal	154489
N conservative	154489
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.	180
Corr. Est. Fragment Len.	0.2092
Phantom Peak	75
Corr. Phantom Peak	0.2102
Argmin. Corr.	1500
Min. Corr.	0.1924
NSC	1.0871
RSC	0.9432

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	0.4468

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	0.1620
AUC	0.4933
CHANCE divergence	0.1625
Elbow Point	0.0000
JS Distance	0.7855
Synthetic AUC	0.4956
Synthetic Elbow Point	0.3569
Synthetic JS Distance	0.4650