QC Report

	general
Report generated at	2021-03-28 00:13:47
Title	elk1_gm12878
Description	chipseq of elk1_gm12878
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	30090669	29164315	5526289	6211790
Total Reads (QC-failed)	0	0	0	0
Duplicate Reads	0	0	0	0
Duplicate Reads (QC-failed)	0	0	0	0
Mapped Reads	29335345	27673782	4962259	5760784
Mapped Reads (QC-failed)	0	0	0	0
% Mapped Reads	97.5	94.89999999999999	89.8	92.7
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	24322162	22602615	3930750	4510303
Paired Reads	0	0	0	0
Unmapped Reads	0	0	0	0
Unpaired Duplicate Reads	1269525	1281845	126940	127662
Paired Duplicate Reads	0	0	0	0
Paired Optical Duplicate Reads	0	0	0	0
% Duplicate Reads	5.2196	5.6712	3.2294000000000005	2.8305000000000002

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	24322162	22602615	3930750	4510303
Total Reads (QC-failed)	0	0	0	0
Duplicate Reads	0	0	0	0
Duplicate Reads (QC-failed)	0	0	0	0
Mapped Reads	24322162	22602615	3930750	4510303
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	24241533	22579184	3898040	4488260
Distinct Fragments	23031914	21307391	3789354	4370528
Positions with Two Read	714676	1132481	101113	110480
NRF = Distinct/Total	0.950101	0.943674	0.972118	0.973769
PBC1 = OneRead/Distinct	0.962267	0.944061	0.972505	0.974007
PBC2 = OneRead/TwoRead	31.01106	17.762316	36.446016	38.531164

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	3269	1294
N1	4791	1400
N2	3320	1233
Np	3524	1534
N optimal	3524	1534
N conservative	3269	1294
Optimal Set	pooled-pr1_vs_pooled-pr2	pooled-pr1_vs_pooled-pr2
Conservative Set	rep1_vs_rep2	rep1_vs_rep2
Rescue Ratio	1.078005506271031	1.1854714064914993
Self Consistency Ratio	1.4430722891566266	1.1354420113544201
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	7094	4879

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	110.0	124.0	115.0
25 percentile	137.0	124.0	208.0	170.75
50 percentile (median)	183.5	165.0	260.0	214.0
75 percentile	260.0	224.0	333.75	281.0
Max size	1038.0	1108.0	1089.0	1089.0
Mean	215.35959966168593	187.609756097561	284.08279009126466	238.433314415437

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	110
Cross-correlation at Estimated Fragment Length	0.191092348722724	0.166042050298507
Phantom Peak	35	35
Cross-correlation at Phantom Peak	0.1907363	0.1640176
Argmin of Cross-correlation	1500	1500
Minimum of Cross-correlation	0.1846639	0.1589394
NSC (Normalized Strand Cross-correlation coeff.)	1.034812	1.044688
RSC (Relative Strand Cross-correlation coeff.)	1.058638	1.398651

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	0.27505174059633974	0.28975906726118494
Synthetic AUC	0.4881482441270383	0.48768785786095875
X-intercept	0.13448135219107257	0.13727442489093622
Synthetic X-intercept	3.7535452756097367e-60	1.1467019902053372e-55
Elbow Point	0.5438914280458887	0.5355761887053498
Synthetic Elbow Point	0.4957437407725736	0.5095954829328775
JS Distance	0.15115577212194892	0.12587483022235707
Synthetic JS Distance	0.26770444599718685	0.22302253903403263
% Genome Enriched	33.99231455156888	43.543143676299465
Diff. Enrichment	16.86450279420253	16.110313242589065
CHANCE Divergence	0.14473312438422709	0.14415022580622625

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	0.010872265384960432	0.008122998157514076	0.02063961254760165	0.018482550869333002	0.020672257671830323	0.018619350841455773	0.008393731951033033	0.008877271619697381	0.008941204109317431

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	0.0073539614263910084	0.009312042243613048	0.006954991712242145	0.007632875911163094

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	0.004550666271679884	0.005150734544075481	0.004282336357983357	0.005037466667129819

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