Untitled

No description

Report generated at 2022-07-12 17:08:57

Pipeline type: Histone ChIP-Seq

Peak caller: MACS2

Alignment

Flagstat (raw BAM)

	rep1	ctl1
Total	49190065	34727622
Total(QC-failed)	0	0
Dupes	0	0
Dupes(QC-failed)	0	0
Mapped	35456158	32997736
Mapped(QC-failed)	0	0
% Mapped	72.0800	95.0200
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	28348988	22695585
Paired Reads	0	0
Unmapped Reads	0	0
Unpaired Dupes	3934586	196955
Paired Dupes	0	0
Paired Opt. Dupes	0	0
% Dupes/100	0.1388	0.0087

Library complexity (filtered non-mito BAM)

	rep1	ctl1
Total Reads	28344841	22680340
Distinct Reads	24763756	22503266
One Read	22151128	22338296
Two Reads	2019400	162449
NRF = Distinct/Total	0.8737	0.9922
PBC1 = OneRead/Distinct	0.8945	0.9927
PBC2 = OneRead/TwoReads	10.9692	137.5096

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	24414402	22498630
Total(QC-failed)	0	0
Dupes	0	0
Dupes(QC-failed)	0	0
Mapped	24414402	22498630
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	48890
Np	0
N optimal	48890
N conservative	48890
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.	175
Corr. Est. Fragment Len.	0.3935
Phantom Peak	40
Corr. Phantom Peak	0.3303
Argmin. Corr.	1500
Min. Corr.	0.1873
NSC	2.1008
RSC	1.4423

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	0.4586

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	0.1311
AUC	0.4881
CHANCE divergence	0.2511
Elbow Point	0.0000
JS Distance	0.8061
Synthetic AUC	0.4931
Synthetic Elbow Point	0.4193
Synthetic JS Distance	0.5198