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Report generated at 2021-12-31 21:29:00

Pipeline type: Histone ChIP-Seq

Peak caller: MACS2

Alignment

Flagstat (raw BAM)

	rep1	ctl1
Total	33242274	36388082
Total(QC-failed)	0	0
Dupes	0	0
Dupes(QC-failed)	0	0
Mapped	32195166	36009049
Mapped(QC-failed)	0	0
% Mapped	96.8500	98.9600
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	21379501	28647216
Paired Reads	0	0
Unmapped Reads	0	0
Unpaired Dupes	660009	5063418
Paired Dupes	0	0
Paired Opt. Dupes	0	0
% Dupes/100	0.0309	0.1768

Library complexity (filtered non-mito BAM)

	rep1	ctl1
Total Reads	21379254	28645171
Distinct Reads	20736558	23761517
One Read	20174681	19678510
Two Reads	541448	3441076
NRF = Distinct/Total	0.9699	0.8295
PBC1 = OneRead/Distinct	0.9729	0.8282
PBC2 = OneRead/TwoReads	37.2606	5.7187

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	20719492	23583798
Total(QC-failed)	0	0
Dupes	0	0
Dupes(QC-failed)	0	0
Mapped	20719492	23583798
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	35060
Np	0
N optimal	35060
N conservative	35060
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.	-5
Corr. Est. Fragment Len.	0.2040
Phantom Peak	40
Corr. Phantom Peak	0.2365
Argmin. Corr.	1500
Min. Corr.	0.1949
NSC	1.0464
RSC	0.2176

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	0.0464

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	0.2357
AUC	0.4881
CHANCE divergence	0.2037
Elbow Point	0.0000
JS Distance	0.6159
Synthetic AUC	0.5216
Synthetic Elbow Point	0.1043
Synthetic JS Distance	0.2805