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Report generated at 2021-06-17 08:36:25

Pipeline type: Histone ChIP-Seq

Peak caller: MACS2

Alignment

Flagstat (raw BAM)

	rep1	ctl1
Total	66248696	75195502
Total(QC-failed)	0	0
Dupes	0	0
Dupes(QC-failed)	0	0
Mapped	63739309	74386272
Mapped(QC-failed)	0	0
% Mapped	96.2100	98.9200
Paired	66248696	75195502
Paired(QC-failed)	0	0
Read1	33124348	37597751
Read1(QC-failed)	0	0
Read2	33124348	37597751
Read2(QC-failed)	0	0
Properly Paired	63359473	72621158
Properly Paired(QC-failed)	0	0
% Properly Paired	95.6400	96.5800
With itself	63479205	74067441
With itself(QC-failed)	0	0
Singletons	260104	318831
Singletons(QC-failed)	0	0
% Singleton	0.3900	0.4200
Diff. Chroms	22955	94964
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	29066057	31937516
Unmapped Reads	0	0
Unpaired Dupes	0	0
Paired Dupes	2275868	278644
Paired Opt. Dupes	43334	70103
% Dupes/100	0.0783	0.0087

Library complexity (filtered non-mito BAM)

	rep1	ctl1
Total Read Pairs	29064682	31937133
Distinct Read Pairs	26788872	31658497
One Read Pair	24935645	31384633
Two Read Pairs	1612447	270131
NRF = Distinct/Total	0.9217	0.9913
PBC1 = OnePair/Distinct	0.9308	0.9913
PBC2 = OnePair/TwoPair	15.4645	116.1830

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	53580378	63317744
Total(QC-failed)	0	0
Dupes	0	0
Dupes(QC-failed)	0	0
Mapped	53580378	63317744
Mapped(QC-failed)	0	0
% Mapped	100.0000	100.0000
Paired	53580378	63317744
Paired(QC-failed)	0	0
Read1	26790189	31658872
Read1(QC-failed)	0	0
Read2	26790189	31658872
Read2(QC-failed)	0	0
Properly Paired	53580378	63317744
Properly Paired(QC-failed)	0	0
% Properly Paired	100.0000	100.0000
With itself	53580378	63317744
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	102569
Np	0
N optimal	102569
N conservative	102569
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.	155
Corr. Est. Fragment Len.	0.3330
Phantom Peak	55
Corr. Phantom Peak	0.1960
Argmin. Corr.	1500
Min. Corr.	0.1344
NSC	2.4777
RSC	3.2260

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	0.6556

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	0.0866
AUC	0.4943
CHANCE divergence	0.2568
Elbow Point	0.0000
JS Distance	0.8934
Synthetic AUC	0.4980
Synthetic Elbow Point	0.5215
Synthetic JS Distance	0.6250