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Report generated at 2022-01-05 22:05:47

Pipeline type: Histone ChIP-Seq

Peak caller: MACS2

Alignment

Flagstat (raw BAM)

	rep1	ctl1
Total	130748586	107365242
Total(QC-failed)	0	0
Dupes	0	0
Dupes(QC-failed)	0	0
Mapped	127979593	105642663
Mapped(QC-failed)	0	0
% Mapped	97.8800	98.4000
Paired	130748586	107365242
Paired(QC-failed)	0	0
Read1	65374293	53682621
Read1(QC-failed)	0	0
Read2	65374293	53682621
Read2(QC-failed)	0	0
Properly Paired	126378375	101883703
Properly Paired(QC-failed)	0	0
% Properly Paired	96.6600	94.8900
With itself	127192301	105057441
With itself(QC-failed)	0	0
Singletons	787292	585222
Singletons(QC-failed)	0	0
% Singleton	0.6000	0.5500
Diff. Chroms	95982	159211
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	53181346	44782756
Unmapped Reads	0	0
Unpaired Dupes	0	0
Paired Dupes	500345	288172
Paired Opt. Dupes	4291	2765
% Dupes/100	0.0094	0.0064

Library complexity (filtered non-mito BAM)

	rep1	ctl1
Total Read Pairs	53134649	44758305
Distinct Read Pairs	52640240	44472498
One Read Pair	52175530	44200745
Two Read Pairs	454380	266773
NRF = Distinct/Total	0.9907	0.9936
PBC1 = OnePair/Distinct	0.9912	0.9939
PBC2 = OnePair/TwoPair	114.8280	165.6867

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	105362002	88989168
Total(QC-failed)	0	0
Dupes	0	0
Dupes(QC-failed)	0	0
Mapped	105362002	88989168
Mapped(QC-failed)	0	0
% Mapped	100.0000	100.0000
Paired	105362002	88989168
Paired(QC-failed)	0	0
Read1	52681001	44494584
Read1(QC-failed)	0	0
Read2	52681001	44494584
Read2(QC-failed)	0	0
Properly Paired	105362002	88989168
Properly Paired(QC-failed)	0	0
% Properly Paired	100.0000	100.0000
With itself	105362002	88989168
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	73842
Np	0
N optimal	73842
N conservative	73842
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.	-10
Corr. Est. Fragment Len.	0.1768
Phantom Peak	50
Corr. Phantom Peak	0.1904
Argmin. Corr.	1500
Min. Corr.	0.1716
NSC	1.0299
RSC	0.2738

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	0.0603

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	0.3141
AUC	0.4960
CHANCE divergence	0.0977
Elbow Point	0.0000
JS Distance	0.5468
Synthetic AUC	0.5012
Synthetic Elbow Point	0.0598
Synthetic JS Distance	0.2260