QC Report

	general
Report generated at	2021-03-26 07:59:38
Title	runx1_k562_2
Description	chipseq of runx1_k562_2
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}, 'ctl2': {'paired_end': False}}
Peak caller	macs2

Alignment quality metrics

SAMstat (raw unfiltered BAM)

	rep1	rep2	ctl1	ctl2
Total Reads	30012916	22880516	44629637	51514306
Total Reads (QC-failed)	0	0	0	0
Duplicate Reads	0	0	0	0
Duplicate Reads (QC-failed)	0	0	0	0
Mapped Reads	28849345	22219529	43435421	50089389
Mapped Reads (QC-failed)	0	0	0	0
% Mapped Reads	96.1	97.1	97.3	97.2
Paired Reads	0	0	0	0
Paired Reads (QC-failed)	0	0	0	0
Read1	0	0	0	0
Read1 (QC-failed)	0	0	0	0
Read2	0	0	0	0
Read2 (QC-failed)	0	0	0	0
Properly Paired Reads	0	0	0	0
Properly Paired Reads (QC-failed)	0	0	0	0
% Properly Paired Reads	0.0	0.0	0.0	0.0
With itself	0	0	0	0
With itself (QC-failed)	0	0	0	0
Singletons	0	0	0	0
Singletons (QC-failed)	0	0	0	0
% Singleton	0.0	0.0	0.0	0.0
Diff. Chroms	0	0	0	0
Diff. Chroms (QC-failed)	0	0	0	0

Marking duplicates (filtered BAM)

	rep1	rep2	ctl1	ctl2
Unpaired Reads	23345048	18691722	35858332	41995892
Paired Reads	0	0	0	0
Unmapped Reads	0	0	0	0
Unpaired Duplicate Reads	875719	557520	1050728	1236726
Paired Duplicate Reads	0	0	0	0
Paired Optical Duplicate Reads	0	0	0	0
% Duplicate Reads	3.7512	2.9827	2.9302	2.9449

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	ctl2
Total Reads	23345048	18691722	35858332	41995892
Total Reads (QC-failed)	0	0	0	0
Duplicate Reads	0	0	0	0
Duplicate Reads (QC-failed)	0	0	0	0
Mapped Reads	23345048	18691722	35858332	41995892
Mapped Reads (QC-failed)	0	0	0	0
% Mapped Reads	100.0	100.0	100.0	100.0
Paired Reads	0	0	0	0
Paired Reads (QC-failed)	0	0	0	0
Read1	0	0	0	0
Read1 (QC-failed)	0	0	0	0
Read2	0	0	0	0
Read2 (QC-failed)	0	0	0	0
Properly Paired Reads	0	0	0	0
Properly Paired Reads (QC-failed)	0	0	0	0
% Properly Paired Reads	0.0	0.0	0.0	0.0
With itself	0	0	0	0
With itself (QC-failed)	0	0	0	0
Singletons	0	0	0	0
Singletons (QC-failed)	0	0	0	0
% Singleton	0.0	0.0	0.0	0.0
Diff. Chroms	0	0	0	0
Diff. Chroms (QC-failed)	0	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	ctl2
Total Fragments	23199619	18522740	35351730	41255400
Distinct Fragments	22447948	18111307	34784593	40736002
Positions with Two Read	699321	389333	519095	481469
NRF = Distinct/Total	0.9676	0.977788	0.983957	0.98741
PBC1 = OneRead/Distinct	0.967785	0.977972	0.984692	0.987919
PBC2 = OneRead/TwoRead	31.065552	45.494096	65.984284	83.585612

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	27249	1788
N1	22754	1123
N2	21952	329
Np	36718	3010
N optimal	36718	3010
N conservative	27249	1788
Optimal Set	pooled-pr1_vs_pooled-pr2	pooled-pr1_vs_pooled-pr2
Conservative Set	rep1_vs_rep2	rep1_vs_rep2
Rescue Ratio	1.3474989907886528	1.6834451901565997
Self Consistency Ratio	1.0365342565597668	3.4133738601823707
Reproducibility Test	pass	borderline

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	215823	182834

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	110.0	75.0	92.0	92.0
25 percentile	110.0	75.0	199.0	135.0
50 percentile (median)	131.0	79.0	271.0	178.0
75 percentile	174.0	114.0	374.0	252.0
Max size	3032.0	1925.0	2999.0	2999.0
Mean	155.1023245900576	101.27295798374482	312.9737541528239	210.89487444849937

Enrichment / Signal-to-noise ratio

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

	rep1	rep2
Number of Subsampled Reads	15000000	15000000
Estimated Fragment Length	110	75
Cross-correlation at Estimated Fragment Length	0.186159364456318	0.184217440345149
Phantom Peak	55	50
Cross-correlation at Phantom Peak	0.184269	0.1838287
Argmin of Cross-correlation	1500	1500
Minimum of Cross-correlation	0.1743143	0.1736365
NSC (Normalized Strand Cross-correlation coeff.)	1.067953	1.060937
RSC (Relative Strand Cross-correlation coeff.)	1.189893	1.038137

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	0.20156882249652308	0.21641091524630007
Synthetic AUC	0.4897288720547548	0.48853027075412603
X-intercept	0.24741387928104444	0.22344202970989863
Synthetic X-intercept	1.9884260907950303e-80	2.7450045460861697e-64
Elbow Point	0.628206409820661	0.6639462582721875
Synthetic Elbow Point	0.49713979607071634	0.5092306120553107
JS Distance	0.0864967013479727	0.06893302370318925
Synthetic JS Distance	0.3328969468639794	0.31393212561347944
% Genome Enriched	36.243277285722854	31.201191594857747
Diff. Enrichment	22.302206725269365	18.62291933141893
CHANCE Divergence	0.20657454198368028	0.16417556672790695

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	0.10446309641342352	0.07453037232203646	0.0913165824289588	0.10886423412460339	0.091202382620931	0.10856709724229796	0.09587715706987002	0.09196448728101612	0.09193194434301208

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	0.025345953078697532	0.0236600927100257	0.017802854119058693	0.029067147642409252

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	0.004914269102978179	0.0035689795968721076	0.0017350996339449089	0.006925841352701456

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