QC Report

	general
Report generated at	2022-06-01 10:31:30
Title	dmd-6_RJP5356_L4larva_1_2
Description	gevirl
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	16022412	15929872	19803610
Total Reads (QC-failed)	0	0	0
Duplicate Reads	0	0	0
Duplicate Reads (QC-failed)	0	0	0
Mapped Reads	15941676	15843442	19677847
Mapped Reads (QC-failed)	0	0	0
% Mapped Reads	99.5	99.5	99.4
Paired Reads	16022412	15929872	19803610
Paired Reads (QC-failed)	0	0	0
Read1	8011206	7964936	9901805
Read1 (QC-failed)	0	0	0
Read2	8011206	7964936	9901805
Read2 (QC-failed)	0	0	0
Properly Paired Reads	15886400	15777976	19587140
Properly Paired Reads (QC-failed)	0	0	0
% Properly Paired Reads	99.2	99.0	98.9
With itself	15930552	15831636	19663322
With itself (QC-failed)	0	0	0
Singletons	11124	11806	14525
Singletons (QC-failed)	0	0	0
% Singleton	0.1	0.1	0.1
Diff. Chroms	3747	4365	14547
Diff. Chroms (QC-failed)	0	0	0

Marking duplicates (filtered BAM)

	rep1	rep2	ctl1
Unpaired Reads	0	0	0
Paired Reads	7373992	7301453	9059600
Unmapped Reads	0	0	0
Unpaired Duplicate Reads	0	0	0
Paired Duplicate Reads	1113185	1179505	1503770
Paired Optical Duplicate Reads	60256	58139	93148
% Duplicate Reads	15.096100000000002	16.1544	16.5986

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	12521614	12243896	15111660
Total Reads (QC-failed)	0	0	0
Duplicate Reads	0	0	0
Duplicate Reads (QC-failed)	0	0	0
Mapped Reads	12521614	12243896	15111660
Mapped Reads (QC-failed)	0	0	0
% Mapped Reads	100.0	100.0	100.0
Paired Reads	12521614	12243896	15111660
Paired Reads (QC-failed)	0	0	0
Read1	6260807	6121948	7555830
Read1 (QC-failed)	0	0	0
Read2	6260807	6121948	7555830
Read2 (QC-failed)	0	0	0
Properly Paired Reads	12521614	12243896	15111660
Properly Paired Reads (QC-failed)	0	0	0
% Properly Paired Reads	100.0	100.0	100.0
With itself	12521614	12243896	15111660
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	7298967	7202823	8800571
Distinct Fragments	6201885	6047211	7355422
Positions with Two Read	845460	863150	1098823
NRF = Distinct/Total	0.849694	0.839561	0.835789
PBC1 = OneRead/Distinct	0.844662	0.834762	0.828501
PBC2 = OneRead/TwoRead	6.196032	5.848322	5.545914

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	25625	6626
N1	20700	6533
N2	23312	4653
Np	25431	7338
N optimal	25625	7338
N conservative	25625	6626
Optimal Set	rep1_vs_rep2	pooled-pr1_vs_pooled-pr2
Conservative Set	rep1_vs_rep2	rep1_vs_rep2
Rescue Ratio	1.0076284849199795	1.107455478418352
Self Consistency Ratio	1.126183574879227	1.404040404040404
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	42865	51190

Top 300000 raw peaks from spp with FDR 0.01

Peak calling statistics

Peak region size

	rep1	rep2	idr_opt	overlap_opt
Min size	84.0	79.0	86.0	86.0
25 percentile	336.0	316.0	222.0	344.0
50 percentile (median)	336.0	316.0	344.0	344.0
75 percentile	336.0	316.0	344.0	344.0
Max size	783.0	656.0	805.0	805.0
Mean	327.3290796687274	312.4155108419613	291.49345870809486	328.91258536585366

Enrichment / Signal-to-noise ratio

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

	rep1	rep2
Number of Subsampled Reads	6927043	6833572
Estimated Fragment Length	170	150
Cross-correlation at Estimated Fragment Length	0.685711943934062	0.675026234671933
Phantom Peak	55	55
Cross-correlation at Phantom Peak	0.6723523	0.6682276
Argmin of Cross-correlation	1500	1500
Minimum of Cross-correlation	0.6490589	0.6530976
NSC (Normalized Strand Cross-correlation coeff.)	1.056471	1.033576
RSC (Relative Strand Cross-correlation coeff.)	1.573535	1.449347

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	0.3550016246506134	0.3713718321395771
Synthetic AUC	0.4981984787023762	0.4981734885248857
X-intercept	0.01916215962610031	0.019220057378669272
Synthetic X-intercept	0.0	0.0
Elbow Point	0.7057316778569047	0.658844480647177
Synthetic Elbow Point	0.5015080061009015	0.50172041146985
JS Distance	0.15848412273761095	0.1221611463414908
Synthetic JS Distance	0.23119122920471707	0.20611435823072574
% Genome Enriched	28.78796046182442	34.56655546305225
Diff. Enrichment	14.558298892506084	12.029716338745033
CHANCE Divergence	0.1258942295951572	0.10269150459364829

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	0.3204539766199469	0.32319532932981465	0.3356977246387367	0.34329497735034664	0.34031145510657895	0.3490585022937144	0.418654532048805	0.31329697524525235	0.3234361273752188

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	0.21856133792520324	0.212963999688858	0.19175203709668884	0.21891877049977973

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	0.10614826829732156	0.11841229093949071	0.07680365792064879	0.11260462635334383

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