QC Report

	general
Report generated at	2021-03-28 09:03:34
Title	gabpa_mhepg2
Description	chipseq of gabpa_mhepg2
Pipeline version	v1.3.6
Pipeline type	tf
Genome	hg38
Aligner	bowtie2
Sequencing endedness	{'rep1': {'paired_end': False}, 'rep2': {'paired_end': False}, 'ctl1': {'paired_end': False}}
Peak caller	macs2

Alignment quality metrics

SAMstat (raw unfiltered BAM)

	rep1	rep2	ctl1
Total Reads	25397372	34741544	36505927
Total Reads (QC-failed)	0	0	0
Duplicate Reads	0	0	0
Duplicate Reads (QC-failed)	0	0	0
Mapped Reads	24542337	33066260	34605694
Mapped Reads (QC-failed)	0	0	0
% Mapped Reads	96.6	95.19999999999999	94.8
Paired Reads	0	0	0
Paired Reads (QC-failed)	0	0	0
Read1	0	0	0
Read1 (QC-failed)	0	0	0
Read2	0	0	0
Read2 (QC-failed)	0	0	0
Properly Paired Reads	0	0	0
Properly Paired Reads (QC-failed)	0	0	0
% Properly Paired Reads	0.0	0.0	0.0
With itself	0	0	0
With itself (QC-failed)	0	0	0
Singletons	0	0	0
Singletons (QC-failed)	0	0	0
% Singleton	0.0	0.0	0.0
Diff. Chroms	0	0	0
Diff. Chroms (QC-failed)	0	0	0

Marking duplicates (filtered BAM)

	rep1	rep2	ctl1
Unpaired Reads	21619953	28972645	28410036
Paired Reads	0	0	0
Unmapped Reads	0	0	0
Unpaired Duplicate Reads	3002545	5393509	1350001
Paired Duplicate Reads	0	0	0
Paired Optical Duplicate Reads	0	0	0
% Duplicate Reads	13.8878	18.6159	4.751799999999999

Filtered out (samtools view -F 1804):

read unmapped (0x4)
mate unmapped (0x8, for paired-end)
not primary alignment (0x100)
read fails platform/vendor quality checks (0x200)
read is PCR or optical duplicate (0x400)

SAMstat (filtered/deduped BAM)

	rep1	rep2	ctl1
Total Reads	21619953	28972645	28410036
Total Reads (QC-failed)	0	0	0
Duplicate Reads	0	0	0
Duplicate Reads (QC-failed)	0	0	0
Mapped Reads	21619953	28972645	28410036
Mapped Reads (QC-failed)	0	0	0
% Mapped Reads	100.0	100.0	100.0
Paired Reads	0	0	0
Paired Reads (QC-failed)	0	0	0
Read1	0	0	0
Read1 (QC-failed)	0	0	0
Read2	0	0	0
Read2 (QC-failed)	0	0	0
Properly Paired Reads	0	0	0
Properly Paired Reads (QC-failed)	0	0	0
% Properly Paired Reads	0.0	0.0	0.0
With itself	0	0	0
With itself (QC-failed)	0	0	0
Singletons	0	0	0
Singletons (QC-failed)	0	0	0
% Singleton	0.0	0.0	0.0
Diff. Chroms	0	0	0
Diff. Chroms (QC-failed)	0	0	0

Filtered and duplicates removed

Sequence quality metrics (filtered/deduped BAM)

Open chromatin assays are known to have significant GC bias. Please take this into consideration as necessary.

Library complexity quality metrics

Library complexity (filtered non-mito BAM)

	rep1	rep2	ctl1
Total Fragments	21424233	28812530	27459469
Distinct Fragments	18596284	23560731	27036169
Positions with Two Read	800513	1289780	388538
NRF = Distinct/Total	0.868002	0.817725	0.984585
PBC1 = OneRead/Distinct	0.933927	0.915378	0.985206
PBC2 = OneRead/TwoRead	21.695564	16.721431	68.554898

Mitochondrial reads are filtered out by default. The non-redundant fraction (NRF) is the fraction of non-redundant mapped reads in a dataset; it is the ratio between the number of positions in the genome that uniquely mapped reads map to and the total number of uniquely mappable reads. The NRF should be > 0.8. The PBC1 is the ratio of genomic locations with EXACTLY one read pair over the genomic locations with AT LEAST one read pair. PBC1 is the primary measure, and the PBC1 should be close to 1. Provisionally 0-0.5 is severe bottlenecking, 0.5-0.8 is moderate bottlenecking, 0.8-0.9 is mild bottlenecking, and 0.9-1.0 is no bottlenecking. The PBC2 is the ratio of genomic locations with EXACTLY one read pair over the genomic locations with EXACTLY two read pairs. The PBC2 should be significantly greater than 1. See more details at the ENCODE portal standard for ChIP-Seq pipeline

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

Replication quality metrics

IDR (Irreproducible Discovery Rate) plots

Reproducibility QC and peak detection statistics

	overlap	idr
Nt	20173	9534
N1	15279	3567
N2	18742	4206
Np	22057	4928
N optimal	22057	9534
N conservative	20173	9534
Optimal Set	pooled-pr1_vs_pooled-pr2	rep1_vs_rep2
Conservative Set	rep1_vs_rep2	rep1_vs_rep2
Rescue Ratio	1.0933921578347296	1.9346590909090908
Self Consistency Ratio	1.2266509588323844	1.1791421362489487
Reproducibility Test	pass	pass

Reproducibility QC

N1: Replicate 1 self-consistent peaks (comparing two pseudoreplicates generated by subsampling Rep1 reads)
N2: Replicate 2 self-consistent peaks (comparing two pseudoreplicates generated by subsampling Rep2 reads)
Ni: Replicate i self-consistent peaks (comparing two pseudoreplicates generated by subsampling RepX reads)
Nt: True Replicate consistent peaks (comparing true replicates Rep1 vs Rep2)
Np: Pooled-pseudoreplicate consistent 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'

Number of raw peaks

	rep1	rep2
Number of peaks	36263	27919

Top 500000 raw peaks from macs2 with p-val threshold 0.01

Peak calling statistics

Peak region size

	rep1	rep2	idr_opt	overlap_opt
Min size	115.0	130.0	122.0	122.0
25 percentile	122.0	166.0	268.0	180.0
50 percentile (median)	166.0	227.0	371.0	241.0
75 percentile	254.0	341.0	574.0	363.0
Max size	3878.0	3424.0	3336.0	3336.0
Mean	229.32515235915395	300.1055195386654	455.00618837843507	316.9574284807544

Enrichment / Signal-to-noise ratio

Strand cross-correlation measures (trimmed/filtered SE BAM)

	rep1	rep2
Number of Subsampled Reads	15000000	15000000
Estimated Fragment Length	115	130
Cross-correlation at Estimated Fragment Length	0.308421744888589	0.323978692056006
Phantom Peak	55	55
Cross-correlation at Phantom Peak	0.2618186	0.2552089
Argmin of Cross-correlation	1500	1500
Minimum of Cross-correlation	0.1277133	0.1178553
NSC (Normalized Strand Cross-correlation coeff.)	2.414954	2.748953
RSC (Relative Strand Cross-correlation coeff.)	1.347511	1.500677

Performed on subsampled (15000000) reads mapped from FASTQs that are trimmed to 50. Such FASTQ trimming and subsampling reads are for cross-corrleation analysis only. Untrimmed FASTQs are used for all the other analyses.

NOTE1: For SE datasets, reads from replicates are randomly subsampled to 15000000.
NOTE2: For PE datasets, the first end (R1) of each read-pair is selected and trimmed to 50 the reads are then randomly subsampled to 15000000.

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

Jensen-Shannon distance (filtered/deduped BAM)

	rep1	rep2
AUC	0.17175312414875082	0.1679409543168293
Synthetic AUC	0.48944691537890117	0.49089563855403046
X-intercept	0.2099994401880968	0.17230610514867004
Synthetic X-intercept	3.9764204103438765e-76	1.148821595128925e-102
Elbow Point	0.7353529214184035	0.7631055965195694
Synthetic Elbow Point	0.4957863391065812	0.5117370247048635
JS Distance	0.27597222211627037	0.31474771639111315
Synthetic JS Distance	0.4495392248412259	0.48025575453685726
% Genome Enriched	24.513963308328403	0.995865389229219
Diff. Enrichment	19.53541664232868	21.687417477574755
CHANCE Divergence	0.1709381458336585	0.22322186555056106

Peak enrichment

Fraction of reads in peaks (FRiP)

FRiP for macs2 raw peaks

	rep1	rep2	rep1-pr1	rep2-pr1	rep1-pr2	rep2-pr2	pooled	pooled-pr1	pooled-pr2
Fraction of Reads in Peaks	0.217642193764251	0.26178880112602765	0.21424337905621815	0.26323746888703226	0.21427013343970422	0.2632372109359436	0.23991359368419862	0.24566683665200048	0.24572382093221704

FRiP for overlap peaks

	rep1_vs_rep2	rep1-pr1_vs_rep1-pr2	rep2-pr1_vs_rep2-pr2	pooled-pr1_vs_pooled-pr2
Fraction of Reads in Peaks	0.23433760013668403	0.20246588880188593	0.25205289334128794	0.23615523757052365

FRiP for IDR peaks

	rep1_vs_rep2	rep1-pr1_vs_rep1-pr2	rep2-pr1_vs_rep2-pr2	pooled-pr1_vs_pooled-pr2
Fraction of Reads in Peaks	0.21581832978808482	0.16511613138104417	0.20937939908489542	0.19803841660789984

For macs2 raw peaks:

repX: Peak from true replicate X
repX-prY: Peak from Yth pseudoreplicates from replicate X
pooled: Peak from pooled true replicates (pool of rep1, rep2, ...)
pooled-pr1: Peak from 1st pooled pseudo replicate (pool of rep1-pr1, rep2-pr1, ...)
pooled-pr2: Peak from 2nd pooled pseudo replicate (pool of rep1-pr2, rep2-pr2, ...)

For overlap/IDR peaks:

repX_vs_repY: Comparing two peaks from true replicates X and Y
repX-pr1_vs_repX-pr2: Comparing two peaks from both pseudoreplicates from replicate X
pooled-pr1_vs_pooled-pr2: Comparing two peaks from 1st and 2nd pooled pseudo replicates