QC Report

	general
Report generated at	2021-10-25 09:49:51
Title	hmg-1.1_RW12311_L4larva_1_1
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	13785818	14882996	13001384
Total Reads (QC-failed)	0	0	0
Duplicate Reads	0	0	0
Duplicate Reads (QC-failed)	0	0	0
Mapped Reads	13744850	14819896	12940326
Mapped Reads (QC-failed)	0	0	0
% Mapped Reads	99.7	99.6	99.5
Paired Reads	13785818	14882996	13001384
Paired Reads (QC-failed)	0	0	0
Read1	6892909	7441498	6500692
Read1 (QC-failed)	0	0	0
Read2	6892909	7441498	6500692
Read2 (QC-failed)	0	0	0
Properly Paired Reads	13698262	14769590	12847922
Properly Paired Reads (QC-failed)	0	0	0
% Properly Paired Reads	99.4	99.2	98.8
With itself	13732256	14805980	12927348
With itself (QC-failed)	0	0	0
Singletons	12594	13916	12978
Singletons (QC-failed)	0	0	0
% Singleton	0.1	0.1	0.1
Diff. Chroms	4758	5228	32522
Diff. Chroms (QC-failed)	0	0	0

Marking duplicates (filtered BAM)

	rep1	rep2	ctl1
Unpaired Reads	0	0	0
Paired Reads	6305987	6796895	5889877
Unmapped Reads	0	0	0
Unpaired Duplicate Reads	0	0	0
Paired Duplicate Reads	777464	906494	661125
Paired Optical Duplicate Reads	78711	92207	75146
% Duplicate Reads	12.328999999999999	13.336899999999998	11.2248

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	11057046	11780802	10457504
Total Reads (QC-failed)	0	0	0
Duplicate Reads	0	0	0
Duplicate Reads (QC-failed)	0	0	0
Mapped Reads	11057046	11780802	10457504
Mapped Reads (QC-failed)	0	0	0
% Mapped Reads	100.0	100.0	100.0
Paired Reads	11057046	11780802	10457504
Paired Reads (QC-failed)	0	0	0
Read1	5528523	5890401	5228752
Read1 (QC-failed)	0	0	0
Read2	5528523	5890401	5228752
Read2 (QC-failed)	0	0	0
Properly Paired Reads	11057046	11780802	10457504
Properly Paired Reads (QC-failed)	0	0	0
% Properly Paired Reads	100.0	100.0	100.0
With itself	11057046	11780802	10457504
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	6281835	6769775	5815788
Distinct Fragments	5512552	5872690	5167951
Positions with Two Read	620914	704934	533915
NRF = Distinct/Total	0.877538	0.867487	0.888607
PBC1 = OneRead/Distinct	0.874631	0.864553	0.886219
PBC2 = OneRead/TwoRead	7.765083	7.202447	8.578026

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	63733	4277
N1	30711	2613
N2	27805	3082
Np	63627	4414
N optimal	63733	4414
N conservative	63733	4277
Optimal Set	rep1_vs_rep2	pooled-pr1_vs_pooled-pr2
Conservative Set	rep1_vs_rep2	rep1_vs_rep2
Rescue Ratio	1.0016659594197432	1.0320317979892448
Self Consistency Ratio	1.104513576694839	1.1794871794871795
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	99425	98067

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	88.0	94.0	94.0
25 percentile	336.0	350.0	376.0	376.0
50 percentile (median)	336.0	350.0	376.0	376.0
75 percentile	336.0	350.0	376.0	376.0
Max size	586.0	652.0	1103.0	1103.0
Mean	335.2860548151873	348.99692047273805	346.7299501585863	373.9369086658403

Enrichment / Signal-to-noise ratio

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

	rep1	rep2
Number of Subsampled Reads	5934126	6395771
Estimated Fragment Length	170	180
Cross-correlation at Estimated Fragment Length	0.65171494793242	0.664574555887181
Phantom Peak	50	55
Cross-correlation at Phantom Peak	0.6494801	0.6615322
Argmin of Cross-correlation	1500	1500
Minimum of Cross-correlation	0.6407925	0.6515814
NSC (Normalized Strand Cross-correlation coeff.)	1.017045	1.019941
RSC (Relative Strand Cross-correlation coeff.)	1.257251	1.305736

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	0.3881607724714924	0.3879905941607196
Synthetic AUC	0.4980811067154335	0.4981415253803041
X-intercept	0.019661026420938765	0.01933770023271502
Synthetic X-intercept	0.0	0.0
Elbow Point	0.5930001876090228	0.5998000167650616
Synthetic Elbow Point	0.5029501254280696	0.5031143576105873
JS Distance	0.09534597616300455	0.09788205156513319
Synthetic JS Distance	0.1698091201692221	0.17199355530848037
% Genome Enriched	37.637762902112	36.79212521106015
Diff. Enrichment	11.917525309218618	11.903707918405871
CHANCE Divergence	0.10159024152977134	0.10161123989508788

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	0.49371947986831205	0.4944760976374953	0.3873220049329622	0.3584173032672473	0.38115503564967274	0.3608763411652859	0.47800313759860386	0.49314121135385236	0.4946122760099421

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	0.3657859094254415	0.2128228461742856	0.20454991094833783	0.3659256336236234

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	0.06015487974173399	0.04124555509672294	0.0488311406982309	0.061270352618162625

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