QC Report

	general
Report generated at	2021-11-05 02:39:01
Title	klf-2_RW12310_midembryonic_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	19524962	18093198	17773936
Total Reads (QC-failed)	0	0	0
Duplicate Reads	0	0	0
Duplicate Reads (QC-failed)	0	0	0
Mapped Reads	19497575	18056045	17750847
Mapped Reads (QC-failed)	0	0	0
% Mapped Reads	99.9	99.8	99.9
Paired Reads	19524962	18093198	17773936
Paired Reads (QC-failed)	0	0	0
Read1	9762481	9046599	8886968
Read1 (QC-failed)	0	0	0
Read2	9762481	9046599	8886968
Read2 (QC-failed)	0	0	0
Properly Paired Reads	19402424	17933862	17679002
Properly Paired Reads (QC-failed)	0	0	0
% Properly Paired Reads	99.4	99.1	99.5
With itself	19486690	18041776	17737642
With itself (QC-failed)	0	0	0
Singletons	10885	14269	13205
Singletons (QC-failed)	0	0	0
% Singleton	0.1	0.1	0.1
Diff. Chroms	9124	7578	10809
Diff. Chroms (QC-failed)	0	0	0

Marking duplicates (filtered BAM)

	rep1	rep2	ctl1
Unpaired Reads	0	0	0
Paired Reads	8721067	7997587	8117143
Unmapped Reads	0	0	0
Unpaired Duplicate Reads	0	0	0
Paired Duplicate Reads	1075352	1023406	971594
Paired Optical Duplicate Reads	109192	94881	101488
% Duplicate Reads	12.3305	12.7964	11.9697

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	15291430	13948362	14291098
Total Reads (QC-failed)	0	0	0
Duplicate Reads	0	0	0
Duplicate Reads (QC-failed)	0	0	0
Mapped Reads	15291430	13948362	14291098
Mapped Reads (QC-failed)	0	0	0
% Mapped Reads	100.0	100.0	100.0
Paired Reads	15291430	13948362	14291098
Paired Reads (QC-failed)	0	0	0
Read1	7645715	6974181	7145549
Read1 (QC-failed)	0	0	0
Read2	7645715	6974181	7145549
Read2 (QC-failed)	0	0	0
Properly Paired Reads	15291430	13948362	14291098
Properly Paired Reads (QC-failed)	0	0	0
% Properly Paired Reads	100.0	100.0	100.0
With itself	15291430	13948362	14291098
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	8692256	7963709	8078724
Distinct Fragments	7623122	6949427	7114947
Positions with Two Read	876996	818406	799577
NRF = Distinct/Total	0.877002	0.872637	0.880702
PBC1 = OneRead/Distinct	0.873081	0.868968	0.876598
PBC2 = OneRead/TwoRead	7.58909	7.378772	7.800307

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	82257	1032
N1	38951	436
N2	30658	456
Np	83491	1027
N optimal	83491	1032
N conservative	82257	1032
Optimal Set	pooled-pr1_vs_pooled-pr2	rep1_vs_rep2
Conservative Set	rep1_vs_rep2	rep1_vs_rep2
Rescue Ratio	1.0150017627679104	1.0048685491723466
Self Consistency Ratio	1.2705003587970514	1.0458715596330275
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	130490	133186

Top 300000 raw peaks from spp with FDR 0.01

Peak calling statistics

Peak region size

	rep1	rep2	idr_opt	overlap_opt
Min size	70.0	69.0	74.0	74.0
25 percentile	280.0	276.0	296.0	296.0
50 percentile (median)	280.0	276.0	296.0	296.0
75 percentile	280.0	276.0	296.0	296.0
Max size	280.0	276.0	311.0	311.0
Mean	279.94974327534675	275.93953568693405	282.6589147286822	295.83162257009735

Enrichment / Signal-to-noise ratio

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

	rep1	rep2
Number of Subsampled Reads	8176224	7506721
Estimated Fragment Length	180	130
Cross-correlation at Estimated Fragment Length	0.719078168707269	0.699801987211403
Phantom Peak	50	50
Cross-correlation at Phantom Peak	0.7196771	0.7003488
Argmin of Cross-correlation	1500	1500
Minimum of Cross-correlation	0.7151981	0.6958198
NSC (Normalized Strand Cross-correlation coeff.)	1.005425	1.005723
RSC (Relative Strand Cross-correlation coeff.)	0.8662809	0.8792727

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	0.4191544922719077	0.41781776075777194
Synthetic AUC	0.49836765774894226	0.49828584171977713
X-intercept	0.01893030405877538	0.019020144143424705
Synthetic X-intercept	0.0	0.0
Elbow Point	0.5034318912336042	0.5030226197368683
Synthetic Elbow Point	0.49850428038352457	0.50276020196518
JS Distance	0.026260996260080816	0.02661189079600799
Synthetic JS Distance	0.12079300711448987	0.12385220181020033
% Genome Enriched	43.38477510032143	43.258599692547264
Diff. Enrichment	7.656437037433666	7.63201601993293
CHANCE Divergence	0.06506110423346123	0.06485372159232017

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	0.5354272295004456	0.5462187603103504	0.4059020764046167	0.3631515495293928	0.40578682383358833	0.3690775116214379	0.5317354514696958	0.5376512202752715	0.539067793514782

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	0.3624897536890823	0.19132435619167076	0.1553334362844899	0.3673447813855858

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	0.00942599044480207	0.0049379292845731235	0.0055136940093754375	0.009411934257261475

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