QC Report

	general
Report generated at	2021-08-27 19:14:20
Title	attf-3_RW12296_youngadult_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	21854230	24380968	23329308
Total Reads (QC-failed)	0	0	0
Duplicate Reads	0	0	0
Duplicate Reads (QC-failed)	0	0	0
Mapped Reads	21808756	24334462	23289640
Mapped Reads (QC-failed)	0	0	0
% Mapped Reads	99.8	99.8	99.8
Paired Reads	21854230	24380968	23329308
Paired Reads (QC-failed)	0	0	0
Read1	10927115	12190484	11664654
Read1 (QC-failed)	0	0	0
Read2	10927115	12190484	11664654
Read2 (QC-failed)	0	0	0
Properly Paired Reads	21695606	24201474	23104426
Properly Paired Reads (QC-failed)	0	0	0
% Properly Paired Reads	99.3	99.3	99.0
With itself	21790092	24316480	23268766
With itself (QC-failed)	0	0	0
Singletons	18664	17982	20874
Singletons (QC-failed)	0	0	0
% Singleton	0.1	0.1	0.1
Diff. Chroms	17638	23742	60380
Diff. Chroms (QC-failed)	0	0	0

Marking duplicates (filtered BAM)

	rep1	rep2	ctl1
Unpaired Reads	0	0	0
Paired Reads	9853429	10900245	10532583
Unmapped Reads	0	0	0
Unpaired Duplicate Reads	0	0	0
Paired Duplicate Reads	1788087	1930047	1911240
Paired Optical Duplicate Reads	176687	176314	203416
% Duplicate Reads	18.1469	17.7065	18.146

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	16130684	17940396	17242686
Total Reads (QC-failed)	0	0	0
Duplicate Reads	0	0	0
Duplicate Reads (QC-failed)	0	0	0
Mapped Reads	16130684	17940396	17242686
Mapped Reads (QC-failed)	0	0	0
% Mapped Reads	100.0	100.0	100.0
Paired Reads	16130684	17940396	17242686
Paired Reads (QC-failed)	0	0	0
Read1	8065342	8970198	8621343
Read1 (QC-failed)	0	0	0
Read2	8065342	8970198	8621343
Read2 (QC-failed)	0	0	0
Properly Paired Reads	16130684	17940396	17242686
Properly Paired Reads (QC-failed)	0	0	0
% Properly Paired Reads	100.0	100.0	100.0
With itself	16130684	17940396	17242686
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	9806852	10842025	10464027
Distinct Fragments	8035049	8932314	8575513
Positions with Two Read	1271300	1378662	1365109
NRF = Distinct/Total	0.81933	0.82386	0.819523
PBC1 = OneRead/Distinct	0.813068	0.818248	0.812593
PBC2 = OneRead/TwoRead	5.138865	5.301405	5.104647

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	45544	1737
N1	29656	964
N2	27805	1063
Np	43766	1700
N optimal	45544	1737
N conservative	45544	1737
Optimal Set	rep1_vs_rep2	rep1_vs_rep2
Conservative Set	rep1_vs_rep2	rep1_vs_rep2
Rescue Ratio	1.0406251428049171	1.021764705882353
Self Consistency Ratio	1.0665707606545585	1.1026970954356847
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	89846	73463

Top 300000 raw peaks from spp with FDR 0.01

Peak calling statistics

Peak region size

	rep1	rep2	idr_opt	overlap_opt
Min size	78.0	75.0	80.0	80.0
25 percentile	310.0	300.0	320.0	320.0
50 percentile (median)	310.0	300.0	320.0	320.0
75 percentile	310.0	300.0	320.0	320.0
Max size	561.0	580.0	862.0	862.0
Mean	309.7297932017007	299.7128078080122	304.475532527346	319.4079132267697

Enrichment / Signal-to-noise ratio

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

	rep1	rep2
Number of Subsampled Reads	9226128	10208160
Estimated Fragment Length	170	165
Cross-correlation at Estimated Fragment Length	0.720177940849619	0.739717401592669
Phantom Peak	50	50
Cross-correlation at Phantom Peak	0.7166533	0.7361429
Argmin of Cross-correlation	1500	1500
Minimum of Cross-correlation	0.7098359	0.7295097
NSC (Normalized Strand Cross-correlation coeff.)	1.01457	1.013992
RSC (Relative Strand Cross-correlation coeff.)	1.517002	1.538877

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	0.41050652335487037	0.41399673721457364
Synthetic AUC	0.4984138232485348	0.4984940575795004
X-intercept	0.01889030444141809	0.018874329553941204
Synthetic X-intercept	0.0	0.0
Elbow Point	0.5337729089870723	0.5297891714225238
Synthetic Elbow Point	0.5003299608996314	0.500620956723687
JS Distance	0.06115277604216892	0.05812000487340814
Synthetic JS Distance	0.14092846006408893	0.1385092084485607
% Genome Enriched	41.78451474282428	41.76794079706701
Diff. Enrichment	8.551074913917123	7.977132944165172
CHANCE Divergence	0.07267755064362576	0.06780081736541156

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	0.4389359434479034	0.3615900117254937	0.3665379595806353	0.34538022460596746	0.3654083112656599	0.3442226135922529	0.5296083364542598	0.39079817839645825	0.3825399723166979

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	0.2418753969642289	0.1752416698510739	0.15879197984258542	0.23456347729511362

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	0.024508938372367416	0.018490164459238057	0.019141996642660506	0.02428167818572232

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