QC Report

	general
Report generated at	2020-11-05 02:15:31
Title	ER fulv
Description	chip seq data for estrogen receptor fulv
Pipeline version	v1.3.6
Pipeline type	tf
Genome	hg19
Aligner	bowtie2
Sequencing endedness	{'rep1': {'paired_end': False}, 'ctl1': {'paired_end': False}}
Peak caller	macs2

Alignment quality metrics

SAMstat (raw unfiltered BAM)

	rep1	ctl1
Total Reads	8302024	3334448
Total Reads (QC-failed)	0	0
Duplicate Reads	0	0
Duplicate Reads (QC-failed)	0	0
Mapped Reads	7593255	2283199
Mapped Reads (QC-failed)	0	0
% Mapped Reads	91.5	68.5
Paired Reads	0	0
Paired Reads (QC-failed)	0	0
Read1	0	0
Read1 (QC-failed)	0	0
Read2	0	0
Read2 (QC-failed)	0	0
Properly Paired Reads	0	0
Properly Paired Reads (QC-failed)	0	0
% Properly Paired Reads	0.0	0.0
With itself	0	0
With itself (QC-failed)	0	0
Singletons	0	0
Singletons (QC-failed)	0	0
% Singleton	0.0	0.0
Diff. Chroms	0	0
Diff. Chroms (QC-failed)	0	0

Marking duplicates (filtered BAM)

	rep1	ctl1
Unpaired Reads	5376404	1401512
Paired Reads	0	0
Unmapped Reads	0	0
Unpaired Duplicate Reads	805801	411572
Paired Duplicate Reads	0	0
Paired Optical Duplicate Reads	0	0
% Duplicate Reads	14.9877	29.366300000000003

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	ctl1
Total Reads	5376404	1401512
Total Reads (QC-failed)	0	0
Duplicate Reads	0	0
Duplicate Reads (QC-failed)	0	0
Mapped Reads	5376404	1401512
Mapped Reads (QC-failed)	0	0
% Mapped Reads	100.0	100.0
Paired Reads	0	0
Paired Reads (QC-failed)	0	0
Read1	0	0
Read1 (QC-failed)	0	0
Read2	0	0
Read2 (QC-failed)	0	0
Properly Paired Reads	0	0
Properly Paired Reads (QC-failed)	0	0
% Properly Paired Reads	0.0	0.0
With itself	0	0
With itself (QC-failed)	0	0
Singletons	0	0
Singletons (QC-failed)	0	0
% Singleton	0.0	0.0
Diff. Chroms	0	0
Diff. Chroms (QC-failed)	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	ctl1
Total Fragments	5355615	1391715
Distinct Fragments	4559558	984761
Positions with Two Read	580709	210255
NRF = Distinct/Total	0.85136	0.707588
PBC1 = OneRead/Distinct	0.852565	0.701121
PBC2 = OneRead/TwoRead	6.694089	3.283808

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	0	0
N1	10778	1453
Np	0	0
N optimal	10778	1453
N conservative	10778	1453
Optimal Set	rep1-pr1_vs_rep1-pr2	rep1-pr1_vs_rep1-pr2
Conservative Set	rep1-pr1_vs_rep1-pr2	rep1-pr1_vs_rep1-pr2
Rescue Ratio	0.0	0.0
Self Consistency Ratio	1.0	1.0
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
Number of peaks	18716

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

Peak calling statistics

Peak region size

	rep1	idr_opt	overlap_opt
Min size	85.0	85.0	85.0
25 percentile	85.0	139.0	85.0
50 percentile (median)	85.0	167.0	97.0
75 percentile	113.0	207.0	140.0
Max size	735.0	735.0	735.0
Mean	106.5363325496901	180.6572608396421	119.54704026721099

Enrichment / Signal-to-noise ratio

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

	rep1
Number of Subsampled Reads	5355615
Estimated Fragment Length	85
Cross-correlation at Estimated Fragment Length	0.107557828213769
Phantom Peak	40
Cross-correlation at Phantom Peak	0.0874991
Argmin of Cross-correlation	1500
Minimum of Cross-correlation	0.0552178
NSC (Normalized Strand Cross-correlation coeff.)	1.947883
RSC (Relative Strand Cross-correlation coeff.)	1.621373

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
AUC	0.13059575354019373
Synthetic AUC	0.47291318057052667
X-intercept	0.5716148411236633
Synthetic X-intercept	4.169100715329775e-11
Elbow Point	0.597630603768076
Synthetic Elbow Point	0.5324629068619062
JS Distance	0.09239210903042445
Synthetic JS Distance	0.24889057172350956
% Genome Enriched	39.23582571898817
Diff. Enrichment	37.79988614248295
CHANCE Divergence	0.422592915864044

Peak enrichment

Fraction of reads in peaks (FRiP)

FRiP for macs2 raw peaks

	rep1	rep1-pr1	rep1-pr2
Fraction of Reads in Peaks	0.05200874041459682	0.16957505425559538	0.16961448581617006

FRiP for overlap peaks

	rep1-pr1_vs_rep1-pr2
Fraction of Reads in Peaks	0.04311171556304177

FRiP for IDR peaks

	rep1-pr1_vs_rep1-pr2
Fraction of Reads in Peaks	0.023296984378406087

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