QC Report

	general
Report generated at	2022-03-21 09:42:33
Title	lag-1_RW12314_youngadult_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	12253786	14088850	11834176
Total Reads (QC-failed)	0	0	0
Duplicate Reads	0	0	0
Duplicate Reads (QC-failed)	0	0	0
Mapped Reads	12187106	13806785	11802244
Mapped Reads (QC-failed)	0	0	0
% Mapped Reads	99.5	98.0	99.7
Paired Reads	12253786	14088850	11834176
Paired Reads (QC-failed)	0	0	0
Read1	6126893	7044425	5917088
Read1 (QC-failed)	0	0	0
Read2	6126893	7044425	5917088
Read2 (QC-failed)	0	0	0
Properly Paired Reads	12129496	13730704	11725524
Properly Paired Reads (QC-failed)	0	0	0
% Properly Paired Reads	99.0	97.5	99.1
With itself	12164358	13776312	11787338
With itself (QC-failed)	0	0	0
Singletons	22748	30473	14906
Singletons (QC-failed)	0	0	0
% Singleton	0.2	0.2	0.1
Diff. Chroms	2877	3093	19187
Diff. Chroms (QC-failed)	0	0	0

Marking duplicates (filtered BAM)

	rep1	rep2	ctl1
Unpaired Reads	0	0	0
Paired Reads	5552944	6278261	5368244
Unmapped Reads	0	0	0
Unpaired Duplicate Reads	0	0	0
Paired Duplicate Reads	702670	823183	549390
Paired Optical Duplicate Reads	72798	87091	74018
% Duplicate Reads	12.654000000000002	13.111600000000001	10.2341

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	9700548	10910156	9637708
Total Reads (QC-failed)	0	0	0
Duplicate Reads	0	0	0
Duplicate Reads (QC-failed)	0	0	0
Mapped Reads	9700548	10910156	9637708
Mapped Reads (QC-failed)	0	0	0
% Mapped Reads	100.0	100.0	100.0
Paired Reads	9700548	10910156	9637708
Paired Reads (QC-failed)	0	0	0
Read1	4850274	5455078	4818854
Read1 (QC-failed)	0	0	0
Read2	4850274	5455078	4818854
Read2 (QC-failed)	0	0	0
Properly Paired Reads	9700548	10910156	9637708
Properly Paired Reads (QC-failed)	0	0	0
% Properly Paired Reads	100.0	100.0	100.0
With itself	9700548	10910156	9637708
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	5504855	6213754	5293616
Distinct Fragments	4819391	5419752	4757176
Positions with Two Read	548762	634759	454442
NRF = Distinct/Total	0.87548	0.872219	0.898663
PBC1 = OneRead/Distinct	0.872746	0.869027	0.896224
PBC2 = OneRead/TwoRead	7.664713	7.420002	9.381822

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	95675	291
N1	41469	47
N2	39015	77
Np	95005	322
N optimal	95675	322
N conservative	95675	291
Optimal Set	rep1_vs_rep2	pooled-pr1_vs_pooled-pr2
Conservative Set	rep1_vs_rep2	rep1_vs_rep2
Rescue Ratio	1.007052260407347	1.1065292096219932
Self Consistency Ratio	1.0628988850442138	1.6382978723404256
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	149458	163263

Top 300000 raw peaks from spp with FDR 0.01

Peak calling statistics

Peak region size

	rep1	rep2	idr_opt	overlap_opt
Min size	75.0	210.0	236.0	236.0
25 percentile	236.0	210.0	236.0	236.0
50 percentile (median)	236.0	210.0	236.0	236.0
75 percentile	236.0	210.0	236.0	236.0
Max size	236.0	210.0	236.0	236.0
Mean	235.99892277429112	210.0	236.0	236.0

Enrichment / Signal-to-noise ratio

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

	rep1	rep2
Number of Subsampled Reads	5180212	5846841
Estimated Fragment Length	145	125
Cross-correlation at Estimated Fragment Length	0.619166999397317	0.646120011609241
Phantom Peak	50	50
Cross-correlation at Phantom Peak	0.6196487	0.6465685
Argmin of Cross-correlation	1500	1500
Minimum of Cross-correlation	0.6159102	0.6425469
NSC (Normalized Strand Cross-correlation coeff.)	1.005288	1.005561
RSC (Relative Strand Cross-correlation coeff.)	0.8711618	0.8884889

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	0.4117334925281126	0.41185850940348073
Synthetic AUC	0.49794581559060963	0.49804579482575995
X-intercept	0.019809053970990892	0.019809053970990892
Synthetic X-intercept	0.0	0.0
Elbow Point	0.4998862466173336	0.5028218821594782
Synthetic Elbow Point	0.4980472167057258	0.5004892826425694
JS Distance	0.02812506123880538	0.02316213166858942
Synthetic JS Distance	0.13485575899516156	0.13725656758659452
% Genome Enriched	45.981312514468634	45.48239416944065
Diff. Enrichment	8.689748806517978	8.564487317230613
CHANCE Divergence	0.07403477172679974	0.07291692954429668

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	0.5716489418948291	0.5703437237744355	0.43690583253647114	0.40158014239209777	0.4383843057113887	0.4107691952342386	0.5741935840716552	0.5670696158656201	0.5664347030552668

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	0.3629555302914447	0.18585651037446543	0.16231041975935082	0.3604427582871502

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	0.0023695939740825934	0.0005789363652445202	0.0008121790375866303	0.0025709456600803158

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