QC Report

	general
Report generated at	2021-03-28 06:30:30
Title	max_l32
Description	chipseq of max_l32
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	68437175	44010907	43319205
Total Reads (QC-failed)	0	0	0
Duplicate Reads	0	0	0
Duplicate Reads (QC-failed)	0	0	0
Mapped Reads	66297737	42115461	42487149
Mapped Reads (QC-failed)	0	0	0
% Mapped Reads	96.89999999999999	95.7	98.1
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	56287889	36005402	35743669
Paired Reads	0	0	0
Unmapped Reads	0	0	0
Unpaired Duplicate Reads	4588616	3114329	1398424
Paired Duplicate Reads	0	0	0
Paired Optical Duplicate Reads	0	0	0
% Duplicate Reads	8.152	8.6496	3.9124

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	56287889	36005402	35743669
Total Reads (QC-failed)	0	0	0
Duplicate Reads	0	0	0
Duplicate Reads (QC-failed)	0	0	0
Mapped Reads	56287889	36005402	35743669
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	55749021	35621111	35383847
Distinct Fragments	51679054	32869381	34322167
Positions with Two Read	3458591	2355603	959853
NRF = Distinct/Total	0.926995	0.92275	0.969995
PBC1 = OneRead/Distinct	0.927774	0.92282	0.971052
PBC2 = OneRead/TwoRead	13.863016	12.876749	34.722603

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	37264	13887
N1	35344	13040
N2	44299	20706
Np	43607	22019
N optimal	43607	22019
N conservative	37264	13887
Optimal Set	pooled-pr1_vs_pooled-pr2	pooled-pr1_vs_pooled-pr2
Conservative Set	rep1_vs_rep2	rep1_vs_rep2
Rescue Ratio	1.1702179046801202	1.585583639374955
Self Consistency Ratio	1.2533669081032142	1.5878834355828222
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	47241	67992

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	135.0	185.0	160.0	160.0
25 percentile	170.0	234.0	305.0	240.0
50 percentile (median)	229.0	317.0	385.0	312.0
75 percentile	321.0	448.0	505.0	422.0
Max size	1752.0	4231.0	3957.0	3957.0
Mean	267.6284794987405	371.3361424873515	429.027249193878	355.4253904189694

Enrichment / Signal-to-noise ratio

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

	rep1	rep2
Number of Subsampled Reads	15000000	15000000
Estimated Fragment Length	135	185
Cross-correlation at Estimated Fragment Length	0.172546487790754	0.178246850029501
Phantom Peak	55	50
Cross-correlation at Phantom Peak	0.1717505	0.1735379
Argmin of Cross-correlation	1500	1500
Minimum of Cross-correlation	0.1646476	0.1610815
NSC (Normalized Strand Cross-correlation coeff.)	1.047975	1.106563
RSC (Relative Strand Cross-correlation coeff.)	1.112072	1.378038

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	0.30168191913411707	0.2674472262686064
Synthetic AUC	0.493383716056149	0.49173835841636804
X-intercept	0.09324104397340083	0.11020741113950168
Synthetic X-intercept	1.3304877836316026e-195	5.958631428070965e-125
Elbow Point	0.5779381245426518	0.6048531887413379
Synthetic Elbow Point	0.4967451492319349	0.4988140016146328
JS Distance	0.06839560187192263	0.1178641743172359
Synthetic JS Distance	0.25450750358451385	0.3010935707943662
% Genome Enriched	31.948393133479684	30.42249973008961
Diff. Enrichment	11.108471148941412	17.260125751075044
CHANCE Divergence	0.0945669865332256	0.14725614251272517

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	0.04975429083865625	0.10148999308492654	0.05993321121115039	0.10519349290975838	0.05998828735588729	0.10515238796667234	0.06723478957966728	0.07026603840287764	0.07015238052517143

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	0.058509843364454306	0.04414391877442766	0.08752045040352556	0.06282921474758116

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	0.034672000156544425	0.02601486795854078	0.061608838584832355	0.045501855600750006

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