QC Report

	general
Report generated at	2022-05-07 12:45:11
Title	vab-7_RW12267_L4larva_1_3
Description	mkudron
Pipeline version	v1.3.6
Pipeline type	tf
Genome	WS245chr
Aligner	bwa
Sequencing endedness	{'rep1': {'paired_end': True}, 'rep2': {'paired_end': True}, 'ctl1': {'paired_end': True}}
Peak caller	spp

Alignment quality metrics

SAMstat (raw unfiltered BAM)

	rep1	rep2	ctl1
Total Reads	24159506	14731296	34929134
Total Reads (QC-failed)	0	0	0
Duplicate Reads	0	0	0
Duplicate Reads (QC-failed)	0	0	0
Mapped Reads	24066698	14673583	34838227
Mapped Reads (QC-failed)	0	0	0
% Mapped Reads	99.6	99.6	99.7
Paired Reads	24159506	14731296	34929134
Paired Reads (QC-failed)	0	0	0
Read1	12079753	7365648	17464567
Read1 (QC-failed)	0	0	0
Read2	12079753	7365648	17464567
Read2 (QC-failed)	0	0	0
Properly Paired Reads	23984852	14634970	34655596
Properly Paired Reads (QC-failed)	0	0	0
% Properly Paired Reads	99.3	99.3	99.2
With itself	24046538	14666676	34804706
With itself (QC-failed)	0	0	0
Singletons	20160	6907	33521
Singletons (QC-failed)	0	0	0
% Singleton	0.1	0.0	0.1
Diff. Chroms	7158	4609	49959
Diff. Chroms (QC-failed)	0	0	0

Marking duplicates (filtered BAM)

	rep1	rep2	ctl1
Unpaired Reads	0	0	0
Paired Reads	11015025	6706364	15903071
Unmapped Reads	0	0	0
Unpaired Duplicate Reads	0	0	0
Paired Duplicate Reads	1201147	992749	1858974
Paired Optical Duplicate Reads	61484	31052	125567
% Duplicate Reads	10.9046	14.803099999999999	11.6894

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	19627756	11427230	28088194
Total Reads (QC-failed)	0	0	0
Duplicate Reads	0	0	0
Duplicate Reads (QC-failed)	0	0	0
Mapped Reads	19627756	11427230	28088194
Mapped Reads (QC-failed)	0	0	0
% Mapped Reads	100.0	100.0	100.0
Paired Reads	19627756	11427230	28088194
Paired Reads (QC-failed)	0	0	0
Read1	9813878	5713615	14044097
Read1 (QC-failed)	0	0	0
Read2	9813878	5713615	14044097
Read2 (QC-failed)	0	0	0
Properly Paired Reads	19627756	11427230	28088194
Properly Paired Reads (QC-failed)	0	0	0
% Properly Paired Reads	100.0	100.0	100.0
With itself	19627756	11427230	28088194
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	10933942	6641300	15725585
Distinct Fragments	9747203	5660508	13901193
Positions with Two Read	975428	753256	1511461
NRF = Distinct/Total	0.891463	0.852319	0.883986
PBC1 = OneRead/Distinct	0.889662	0.848112	0.880545
PBC2 = OneRead/TwoRead	8.890164	6.373326	8.09854

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	59275	1732
N1	45156	1014
N2	31975	1119
Np	64426	1815
N optimal	64426	1815
N conservative	59275	1732
Optimal Set	pooled-pr1_vs_pooled-pr2	pooled-pr1_vs_pooled-pr2
Conservative Set	rep1_vs_rep2	rep1_vs_rep2
Rescue Ratio	1.0869000421762969	1.047921478060046
Self Consistency Ratio	1.4122283033620016	1.1035502958579881
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	111593	99174

Top 300000 raw peaks from spp with FDR 0.01

Peak calling statistics

Peak region size

	rep1	rep2	idr_opt	overlap_opt
Min size	72.0	66.0	76.0	76.0
25 percentile	290.0	264.0	304.0	304.0
50 percentile (median)	290.0	264.0	304.0	304.0
75 percentile	290.0	264.0	304.0	304.0
Max size	331.0	315.0	509.0	509.0
Mean	289.8180531036893	263.7362917700204	274.03250688705236	303.15572905348773

Enrichment / Signal-to-noise ratio

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

	rep1	rep2
Number of Subsampled Reads	10288235	6255277
Estimated Fragment Length	180	165
Cross-correlation at Estimated Fragment Length	0.770052557810061	0.652885630444643
Phantom Peak	50	50
Cross-correlation at Phantom Peak	0.769862	0.6524024
Argmin of Cross-correlation	1500	1500
Minimum of Cross-correlation	0.7652323	0.6474769
NSC (Normalized Strand Cross-correlation coeff.)	1.006299	1.008354
RSC (Relative Strand Cross-correlation coeff.)	1.041158	1.098105

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	0.4181406821430364	0.40435667363845557
Synthetic AUC	0.49856271291055243	0.49811423970584634
X-intercept	0.01878664114358333	0.01941406140725969
Synthetic X-intercept	0.0	0.0
Elbow Point	0.5267452873544844	0.5398571719730969
Synthetic Elbow Point	0.5005742819939301	0.5026075788800729
JS Distance	0.04558942948494388	0.062481564933501016
Synthetic JS Distance	0.12958721060254844	0.14792895251402052
% Genome Enriched	42.82143299591178	42.49753228017312
Diff. Enrichment	7.972236515239595	10.021896010047653
CHANCE Divergence	0.06774805754247143	0.08519035916615367

Peak enrichment

Fraction of reads in peaks (FRiP)

FRiP for spp raw peaks

	rep1	rep2	rep1-pr1	rep2-pr1	rep1-pr2	rep2-pr2	pooled	pooled-pr1	pooled-pr2
Fraction of Reads in Peaks	0.48057449868441404	0.4441229414302504	0.43003927703197453	0.4057180251525479	0.4400790390913765	0.40402064262654075	0.44759830192807043	0.4834817517881507	0.479894563784029

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	0.2879220103335419	0.22757344242510452	0.17228435937668185	0.3086031338091732

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	0.016726396205749377	0.010528457761549512	0.01366892939058722	0.017430405539387458

For spp 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