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Report generated at 2021-10-10 05:39:06

Pipeline type: Histone ChIP-Seq

Peak caller: MACS2

Alignment

Flagstat (raw BAM)

	rep1	ctl1
Total	143654712	116616760
Total(QC-failed)	0	0
Dupes	0	0
Dupes(QC-failed)	0	0
Mapped	141998806	115340967
Mapped(QC-failed)	0	0
% Mapped	98.8500	98.9100
Paired	143654712	116616760
Paired(QC-failed)	0	0
Read1	71827356	58308380
Read1(QC-failed)	0	0
Read2	71827356	58308380
Read2(QC-failed)	0	0
Properly Paired	140764449	112618961
Properly Paired(QC-failed)	0	0
% Properly Paired	97.9900	96.5700
With itself	141288392	114670631
With itself(QC-failed)	0	0
Singletons	710414	670336
Singletons(QC-failed)	0	0
% Singleton	0.4900	0.5700
Diff. Chroms	250966	1356484
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	61316856	46805405
Unmapped Reads	0	0
Unpaired Dupes	0	0
Paired Dupes	8133038	2581950
Paired Opt. Dupes	57569	44282
% Dupes/100	0.1326	0.0552

Library complexity (filtered non-mito BAM)

	rep1	ctl1
Total Read Pairs	61316001	46788318
Distinct Read Pairs	53183057	44207269
One Read Pair	45931855	41662261
Two Read Pairs	6457123	2510205
NRF = Distinct/Total	0.8674	0.9448
PBC1 = OnePair/Distinct	0.8637	0.9424
PBC2 = OnePair/TwoPair	7.1134	16.5972

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	106367636	88446910
Total(QC-failed)	0	0
Dupes	0	0
Dupes(QC-failed)	0	0
Mapped	106367636	88446910
Mapped(QC-failed)	0	0
% Mapped	100.0000	100.0000
Paired	106367636	88446910
Paired(QC-failed)	0	0
Read1	53183818	44223455
Read1(QC-failed)	0	0
Read2	53183818	44223455
Read2(QC-failed)	0	0
Properly Paired	106367636	88446910
Properly Paired(QC-failed)	0	0
% Properly Paired	100.0000	100.0000
With itself	106367636	88446910
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	131441
Np	0
N optimal	131441
N conservative	131441
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.	230
Corr. Est. Fragment Len.	0.2055
Phantom Peak	50
Corr. Phantom Peak	0.2063
Argmin. Corr.	1500
Min. Corr.	0.1863
NSC	1.1034
RSC	0.9642

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	0.6077

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	0.1401
AUC	0.4952
CHANCE divergence	0.1251
Elbow Point	0.0000
JS Distance	0.8449
Synthetic AUC	0.5027
Synthetic Elbow Point	0.4330
Synthetic JS Distance	0.5317