QC Report

	general
Report generated at	2021-09-14 14:32:45
Title	tbx-37_RW12282_earlyembryonic_1_2
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	14004780	22451484	21066972
Total Reads (QC-failed)	0	0	0
Duplicate Reads	0	0	0
Duplicate Reads (QC-failed)	0	0	0
Mapped Reads	13943086	22371703	21035061
Mapped Reads (QC-failed)	0	0	0
% Mapped Reads	99.6	99.6	99.8
Paired Reads	14004780	22451484	21066972
Paired Reads (QC-failed)	0	0	0
Read1	7002390	11225742	10533486
Read1 (QC-failed)	0	0	0
Read2	7002390	11225742	10533486
Read2 (QC-failed)	0	0	0
Properly Paired Reads	13795050	22188330	20698658
Properly Paired Reads (QC-failed)	0	0	0
% Properly Paired Reads	98.5	98.8	98.3
With itself	13930604	22344546	21015754
With itself (QC-failed)	0	0	0
Singletons	12482	27157	19307
Singletons (QC-failed)	0	0	0
% Singleton	0.1	0.1	0.1
Diff. Chroms	5638	9100	13075
Diff. Chroms (QC-failed)	0	0	0

Marking duplicates (filtered BAM)

	rep1	rep2	ctl1
Unpaired Reads	0	0	0
Paired Reads	6200845	9983999	9480432
Unmapped Reads	0	0	0
Unpaired Duplicate Reads	0	0	0
Paired Duplicate Reads	789410	1336923	1376128
Paired Optical Duplicate Reads	71790	132178	163096
% Duplicate Reads	12.7307	13.390699999999999	14.515500000000001

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	10822870	17294152	16208608
Total Reads (QC-failed)	0	0	0
Duplicate Reads	0	0	0
Duplicate Reads (QC-failed)	0	0	0
Mapped Reads	10822870	17294152	16208608
Mapped Reads (QC-failed)	0	0	0
% Mapped Reads	100.0	100.0	100.0
Paired Reads	10822870	17294152	16208608
Paired Reads (QC-failed)	0	0	0
Read1	5411435	8647076	8104304
Read1 (QC-failed)	0	0	0
Read2	5411435	8647076	8104304
Read2 (QC-failed)	0	0	0
Properly Paired Reads	10822870	17294152	16208608
Properly Paired Reads (QC-failed)	0	0	0
% Properly Paired Reads	100.0	100.0	100.0
With itself	10822870	17294152	16208608
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	6169340	9932445	9422046
Distinct Fragments	5389934	8615069	8060156
Positions with Two Read	628174	1051458	1075637
NRF = Distinct/Total	0.873665	0.867366	0.855457
PBC1 = OneRead/Distinct	0.870216	0.863447	0.849737
PBC2 = OneRead/TwoRead	7.466731	7.074613	6.367401

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	46821	1107
N1	29637	538
N2	29840	817
Np	29236	1042
N optimal	46821	1107
N conservative	46821	1107
Optimal Set	rep1_vs_rep2	rep1_vs_rep2
Conservative Set	rep1_vs_rep2	rep1_vs_rep2
Rescue Ratio	1.6014844711998906	1.062380038387716
Self Consistency Ratio	1.0068495461753888	1.5185873605947955
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	69581	138367

Top 300000 raw peaks from spp with FDR 0.01

Peak calling statistics

Peak region size

	rep1	rep2	idr_opt	overlap_opt
Min size	59.0	60.0	64.0	64.0
25 percentile	236.0	240.0	256.0	256.0
50 percentile (median)	236.0	240.0	256.0	256.0
75 percentile	236.0	240.0	256.0	256.0
Max size	236.0	251.0	263.0	263.0
Mean	235.8595162472514	239.86961486481604	233.4579945799458	255.47433843788045

Enrichment / Signal-to-noise ratio

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

	rep1	rep2
Number of Subsampled Reads	5826628	9367514
Estimated Fragment Length	140	145
Cross-correlation at Estimated Fragment Length	0.645019504795582	0.742703008843036
Phantom Peak	50	50
Cross-correlation at Phantom Peak	0.6449453	0.742516
Argmin of Cross-correlation	1500	1500
Minimum of Cross-correlation	0.640741	0.7375426
NSC (Normalized Strand Cross-correlation coeff.)	1.006677	1.006997
RSC (Relative Strand Cross-correlation coeff.)	1.017656	1.0376

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	0.4106597921189301	0.41781758937418423
Synthetic AUC	0.49804598545430867	0.49845051037285315
X-intercept	0.019493768941738307	0.019176326370892713
Synthetic X-intercept	0.0	0.0
Elbow Point	0.5119869509141947	0.5098566916494636
Synthetic Elbow Point	0.49935100891499395	0.500777347814283
JS Distance	0.03871764424664884	0.031060458129361625
Synthetic JS Distance	0.13319313565820803	0.12641053177202727
% Genome Enriched	43.28658875015473	41.71914118807374
Diff. Enrichment	8.373278124638261	7.146074191787494
CHANCE Divergence	0.07117348536154246	0.06073263681236188

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	0.3081709380229089	0.5197543076989262	0.3930548194601211	0.3255714417220341	0.38494509957988954	0.3426284214455846	0.5252508960586224	0.2905278311104333	0.28202021409096695

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	0.20411155206977466	0.14229035366774248	0.1381066270262919	0.13676708009831198

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	0.010688756440849248	0.0064541106009773745	0.008659054228273233	0.010303082595304724

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