QC Report

	general
Report generated at	2021-03-27 16:31:59
Title	max_mhepg2
Description	chipseq of max_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	45803823	42887010	51384668
Total Reads (QC-failed)	0	0	0
Duplicate Reads	0	0	0
Duplicate Reads (QC-failed)	0	0	0
Mapped Reads	39508961	38868897	42310468
Mapped Reads (QC-failed)	0	0	0
% Mapped Reads	86.3	90.60000000000001	82.3
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	32070184	31865967	30696132
Paired Reads	0	0	0
Unmapped Reads	0	0	0
Unpaired Duplicate Reads	5904751	6141171	4748982
Paired Duplicate Reads	0	0	0
Paired Optical Duplicate Reads	0	0	0
% Duplicate Reads	18.412	19.2719	15.470900000000002

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	32070184	31865967	30696132
Total Reads (QC-failed)	0	0	0
Duplicate Reads	0	0	0
Duplicate Reads (QC-failed)	0	0	0
Mapped Reads	32070184	31865967	30696132
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	32028840	31821236	30446581
Distinct Fragments	26176411	25735665	25933961
Positions with Two Read	3518864	3601019	3203810
NRF = Distinct/Total	0.817276	0.808758	0.851786
PBC1 = OneRead/Distinct	0.829269	0.82114	0.853756
PBC2 = OneRead/TwoRead	6.168837	5.868498	6.910923

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	67922	32445
N1	71267	34743
N2	63865	31214
Np	71205	33754
N optimal	71205	33754
N conservative	67922	32445
Optimal Set	pooled-pr1_vs_pooled-pr2	pooled-pr1_vs_pooled-pr2
Conservative Set	rep1_vs_rep2	rep1_vs_rep2
Rescue Ratio	1.0483348546862579	1.0403451995685005
Self Consistency Ratio	1.1159007280983324	1.1130582430960467
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	97649	94106

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	120.0	125.0	122.0	122.0
25 percentile	175.0	166.0	411.0	231.0
50 percentile (median)	288.0	268.0	659.0	368.0
75 percentile	543.0	521.0	1083.0	677.0
Max size	6065.0	6333.0	6460.0	6460.0
Mean	445.775635183156	439.8686693728349	819.1892516442496	545.341001334176

Enrichment / Signal-to-noise ratio

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

	rep1	rep2
Number of Subsampled Reads	15000000	15000000
Estimated Fragment Length	120	125
Cross-correlation at Estimated Fragment Length	0.326094588863175	0.32039053998107
Phantom Peak	55	55
Cross-correlation at Phantom Peak	0.2970874	0.2922347
Argmin of Cross-correlation	1500	1500
Minimum of Cross-correlation	0.1483534	0.1475809
NSC (Normalized Strand Cross-correlation coeff.)	2.198093	2.170948
RSC (Relative Strand Cross-correlation coeff.)	1.195027	1.194643

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	0.14660618475580284	0.1474125675420301
Synthetic AUC	0.49382542744843094	0.49376846305420435
X-intercept	0.1848106428665683	0.20786498882371052
Synthetic X-intercept	7.986874739715e-225	1.0829041029423183e-220
Elbow Point	0.8036072024088002	0.7776018361903848
Synthetic Elbow Point	0.5070302249176304	0.49887114076754674
JS Distance	0.3388945667851617	0.31859628960406067
Synthetic JS Distance	0.5102677549234408	0.4966239185111839
% Genome Enriched	8.400810930770984	11.487186254163618
Diff. Enrichment	35.24662049561147	33.44549557418181
CHANCE Divergence	0.3106193769017679	0.2896027500586853

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	0.396338948351528	0.3674272618182276	0.4006725374572219	0.3743435002508005	0.4004356819405838	0.3741647750455768	0.37509998060408734	0.37851261364618877	0.37818329692982766

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	0.3645789844308895	0.3796904002795868	0.3486723939681479	0.3673602591435321

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	0.31344065425521156	0.3276713036632406	0.3027121693812085	0.31760842156419455

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