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Report generated at 2022-07-13 13:48:28

Pipeline type: Histone ChIP-Seq

Peak caller: MACS2

Alignment

Flagstat (raw BAM)

	rep1	ctl1
Total	44787447	51837243
Total(QC-failed)	0	0
Dupes	0	0
Dupes(QC-failed)	0	0
Mapped	40336118	45119622
Mapped(QC-failed)	0	0
% Mapped	90.0600	87.0400
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	32034540	30545616
Paired Reads	0	0
Unmapped Reads	0	0
Unpaired Dupes	3597462	388804
Paired Dupes	0	0
Paired Opt. Dupes	0	0
% Dupes/100	0.1123	0.0127

Library complexity (filtered non-mito BAM)

	rep1	ctl1
Total Reads	32027171	30512152
Distinct Reads	28911116	30212355
One Read	26218605	29928598
Two Reads	2349355	279300
NRF = Distinct/Total	0.9027	0.9902
PBC1 = OneRead/Distinct	0.9069	0.9906
PBC2 = OneRead/TwoReads	11.1599	107.1557

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	28437078	30156812
Total(QC-failed)	0	0
Dupes	0	0
Dupes(QC-failed)	0	0
Mapped	28437078	30156812
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	100955
Np	0
N optimal	100955
N conservative	100955
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.	0.2830
Phantom Peak	40
Corr. Phantom Peak	0.2526
Argmin. Corr.	1500
Min. Corr.	0.1836
NSC	1.5413
RSC	1.4412

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	0.4340

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	0.1100
AUC	0.4890
CHANCE divergence	0.3383
Elbow Point	0.0000
JS Distance	0.7948
Synthetic AUC	0.4965
Synthetic Elbow Point	0.3540
Synthetic JS Distance	0.5110