QC Report

	general
Report generated at	2021-08-31 02:29:53
Title	mdt-29_RJP4521_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	17421290	23714360	26372018
Total Reads (QC-failed)	0	0	0
Duplicate Reads	0	0	0
Duplicate Reads (QC-failed)	0	0	0
Mapped Reads	17366436	23644231	26287816
Mapped Reads (QC-failed)	0	0	0
% Mapped Reads	99.7	99.7	99.7
Paired Reads	17421290	23714360	26372018
Paired Reads (QC-failed)	0	0	0
Read1	8710645	11857180	13186009
Read1 (QC-failed)	0	0	0
Read2	8710645	11857180	13186009
Read2 (QC-failed)	0	0	0
Properly Paired Reads	17296078	23553938	26092648
Properly Paired Reads (QC-failed)	0	0	0
% Properly Paired Reads	99.3	99.3	98.9
With itself	17352750	23625000	26265258
With itself (QC-failed)	0	0	0
Singletons	13686	19231	22558
Singletons (QC-failed)	0	0	0
% Singleton	0.1	0.1	0.1
Diff. Chroms	5429	6772	37940
Diff. Chroms (QC-failed)	0	0	0

Marking duplicates (filtered BAM)

	rep1	rep2	ctl1
Unpaired Reads	0	0	0
Paired Reads	7893575	10757870	11985263
Unmapped Reads	0	0	0
Unpaired Duplicate Reads	0	0	0
Paired Duplicate Reads	1339154	2249330	2429411
Paired Optical Duplicate Reads	121695	209474	257306
% Duplicate Reads	16.9651	20.9087	20.27

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	13108842	17017080	19111704
Total Reads (QC-failed)	0	0	0
Duplicate Reads	0	0	0
Duplicate Reads (QC-failed)	0	0	0
Mapped Reads	13108842	17017080	19111704
Mapped Reads (QC-failed)	0	0	0
% Mapped Reads	100.0	100.0	100.0
Paired Reads	13108842	17017080	19111704
Paired Reads (QC-failed)	0	0	0
Read1	6554421	8508540	9555852
Read1 (QC-failed)	0	0	0
Read2	6554421	8508540	9555852
Read2 (QC-failed)	0	0	0
Properly Paired Reads	13108842	17017080	19111704
Properly Paired Reads (QC-failed)	0	0	0
% Properly Paired Reads	100.0	100.0	100.0
With itself	13108842	17017080	19111704
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	7828164	10653507	11743636
Distinct Fragments	6508245	8438186	9384593
Positions with Two Read	960099	1477992	1634639
NRF = Distinct/Total	0.831388	0.792057	0.799122
PBC1 = OneRead/Distinct	0.826932	0.785255	0.790398
PBC2 = OneRead/TwoRead	5.60554	4.483197	4.537738

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	72746	1690
N1	43343	777
N2	48935	1050
Np	72925	1705
N optimal	72925	1705
N conservative	72746	1690
Optimal Set	pooled-pr1_vs_pooled-pr2	pooled-pr1_vs_pooled-pr2
Conservative Set	rep1_vs_rep2	rep1_vs_rep2
Rescue Ratio	1.0024606163912793	1.0088757396449703
Self Consistency Ratio	1.129017373047551	1.3513513513513513
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	117421	121294

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	71.0	80.0	80.0
25 percentile	290.0	284.0	320.0	320.0
50 percentile (median)	290.0	284.0	320.0	320.0
75 percentile	290.0	284.0	320.0	320.0
Max size	585.0	583.0	818.0	818.0
Mean	289.8976929169399	283.8792685540917	306.76363636363635	319.68769283510454

Enrichment / Signal-to-noise ratio

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

	rep1	rep2
Number of Subsampled Reads	7379147	10049884
Estimated Fragment Length	185	170
Cross-correlation at Estimated Fragment Length	0.679805752027671	0.727700946166669
Phantom Peak	50	50
Cross-correlation at Phantom Peak	0.6796578	0.7275187
Argmin of Cross-correlation	1500	1500
Minimum of Cross-correlation	0.6749419	0.7228518
NSC (Normalized Strand Cross-correlation coeff.)	1.007206	1.006708
RSC (Relative Strand Cross-correlation coeff.)	1.031377	1.039047

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	0.40826775204826266	0.4121004470564197
Synthetic AUC	0.4982406682327819	0.49845449009486426
X-intercept	0.018760856977417487	0.018611105564762493
Synthetic X-intercept	0.0	0.0
Elbow Point	0.5434818201785037	0.5516981810195076
Synthetic Elbow Point	0.49957807913087005	0.5004908173940086
JS Distance	0.06349349695244237	0.058077858984376944
Synthetic JS Distance	0.14399085977934675	0.14262809153715064
% Genome Enriched	43.97040912085937	42.167801449593675
Diff. Enrichment	8.772793390602708	7.784872818092664
CHANCE Divergence	0.0746010050268362	0.06615292946492726

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	0.5126822796399559	0.5081415848077343	0.4534346430547194	0.44946547821365357	0.4566303349495455	0.45640438900210845	0.4883720737244158	0.5093578540528748	0.5112066287104261

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	0.34863812632854857	0.226457302635885	0.24085783224854088	0.34927482053495323

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	0.0189082345761899	0.010706208832176023	0.013011809311585772	0.018997426867134556

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