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Report generated at 2021-10-09 23:14:37

Pipeline type: Histone ChIP-Seq

Peak caller: MACS2

Alignment

Flagstat (raw BAM)

	rep1	ctl1
Total	98148856	135940090
Total(QC-failed)	0	0
Dupes	0	0
Dupes(QC-failed)	0	0
Mapped	96956246	134333469
Mapped(QC-failed)	0	0
% Mapped	98.7800	98.8200
Paired	98148856	135940090
Paired(QC-failed)	0	0
Read1	49074428	67970045
Read1(QC-failed)	0	0
Read2	49074428	67970045
Read2(QC-failed)	0	0
Properly Paired	95390806	131959627
Properly Paired(QC-failed)	0	0
% Properly Paired	97.1900	97.0700
With itself	96319970	133529071
With itself(QC-failed)	0	0
Singletons	636276	804398
Singletons(QC-failed)	0	0
% Singleton	0.6500	0.5900
Diff. Chroms	433125	898860
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	38210123	55193949
Unmapped Reads	0	0
Unpaired Dupes	0	0
Paired Dupes	2220766	3483873
Paired Opt. Dupes	43820	49939
% Dupes/100	0.0581	0.0631

Library complexity (filtered non-mito BAM)

	rep1	ctl1
Total Read Pairs	38209922	55175675
Distinct Read Pairs	35989165	51692963
One Read Pair	33823526	48296105
Two Read Pairs	2112436	3315273
NRF = Distinct/Total	0.9419	0.9369
PBC1 = OnePair/Distinct	0.9398	0.9343
PBC2 = OnePair/TwoPair	16.0116	14.5678

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	71978714	103420152
Total(QC-failed)	0	0
Dupes	0	0
Dupes(QC-failed)	0	0
Mapped	71978714	103420152
Mapped(QC-failed)	0	0
% Mapped	100.0000	100.0000
Paired	71978714	103420152
Paired(QC-failed)	0	0
Read1	35989357	51710076
Read1(QC-failed)	0	0
Read2	35989357	51710076
Read2(QC-failed)	0	0
Properly Paired	71978714	103420152
Properly Paired(QC-failed)	0	0
% Properly Paired	100.0000	100.0000
With itself	71978714	103420152
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	191726
Np	0
N optimal	191726
N conservative	191726
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.	220
Corr. Est. Fragment Len.	0.1777
Phantom Peak	50
Corr. Phantom Peak	0.1876
Argmin. Corr.	1500
Min. Corr.	0.1739
NSC	1.0217
RSC	0.2756

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	0.2141

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	0.2108
AUC	0.4941
CHANCE divergence	0.2088
Elbow Point	0.0000
JS Distance	0.5952
Synthetic AUC	0.4965
Synthetic Elbow Point	0.2157
Synthetic JS Distance	0.3582