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Report generated at 2021-02-06 07:23:57

Pipeline type: Histone ChIP-Seq

Peak caller: MACS2

Alignment

Flagstat (raw BAM)

	rep1	ctl1
Total	58776250	100610396
Total(QC-failed)	0	0
Dupes	0	0
Dupes(QC-failed)	0	0
Mapped	56892045	98910281
Mapped(QC-failed)	0	0
% Mapped	96.7900	98.3100
Paired	58776250	100610396
Paired(QC-failed)	0	0
Read1	29388125	50305198
Read1(QC-failed)	0	0
Read2	29388125	50305198
Read2(QC-failed)	0	0
Properly Paired	56080709	96866538
Properly Paired(QC-failed)	0	0
% Properly Paired	95.4100	96.2800
With itself	56536067	98283644
With itself(QC-failed)	0	0
Singletons	355978	626637
Singletons(QC-failed)	0	0
% Singleton	0.6100	0.6200
Diff. Chroms	37083	146099
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	25695562	42057260
Unmapped Reads	0	0
Unpaired Dupes	0	0
Paired Dupes	2578242	223838
Paired Opt. Dupes	1609	11806
% Dupes/100	0.1003	0.0053

Library complexity (filtered non-mito BAM)

	rep1	ctl1
Total Read Pairs	25677062	42020997
Distinct Read Pairs	23101043	41800825
One Read Pair	20824080	41591190
Two Read Pairs	2047849	205559
NRF = Distinct/Total	0.8997	0.9948
PBC1 = OnePair/Distinct	0.9014	0.9950
PBC2 = OnePair/TwoPair	10.1688	202.3321

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	46234640	83666844
Total(QC-failed)	0	0
Dupes	0	0
Dupes(QC-failed)	0	0
Mapped	46234640	83666844
Mapped(QC-failed)	0	0
% Mapped	100.0000	100.0000
Paired	46234640	83666844
Paired(QC-failed)	0	0
Read1	23117320	41833422
Read1(QC-failed)	0	0
Read2	23117320	41833422
Read2(QC-failed)	0	0
Properly Paired	46234640	83666844
Properly Paired(QC-failed)	0	0
% Properly Paired	100.0000	100.0000
With itself	46234640	83666844
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	80405
Np	0
N optimal	80405
N conservative	80405
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.	145
Corr. Est. Fragment Len.	0.2755
Phantom Peak	55
Corr. Phantom Peak	0.2068
Argmin. Corr.	1500
Min. Corr.	0.1482
NSC	1.8592
RSC	2.1714

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	0.4341

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	0.1610
AUC	0.4938
CHANCE divergence	0.1724
Elbow Point	0.0000
JS Distance	0.8007
Synthetic AUC	0.4944
Synthetic Elbow Point	0.3787
Synthetic JS Distance	0.4911