QC Report

	general
Report generated at	2021-08-29 13:48:52
Title	ekl-4_RW12207_youngadult_1_4
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	12310792	23527318	12345226
Total Reads (QC-failed)	0	0	0
Duplicate Reads	0	0	0
Duplicate Reads (QC-failed)	0	0	0
Mapped Reads	11367851	23344827	12312238
Mapped Reads (QC-failed)	0	0	0
% Mapped Reads	92.30000000000001	99.2	99.7
Paired Reads	12310792	23527318	12345226
Paired Reads (QC-failed)	0	0	0
Read1	6155396	11763659	6172613
Read1 (QC-failed)	0	0	0
Read2	6155396	11763659	6172613
Read2 (QC-failed)	0	0	0
Properly Paired Reads	11315708	23290596	12226386
Properly Paired Reads (QC-failed)	0	0	0
% Properly Paired Reads	91.9	99.0	99.0
With itself	11358650	23333638	12303412
With itself (QC-failed)	0	0	0
Singletons	9201	11189	8826
Singletons (QC-failed)	0	0	0
% Singleton	0.1	0.0	0.1
Diff. Chroms	3954	4681	35477
Diff. Chroms (QC-failed)	0	0	0

Marking duplicates (filtered BAM)

	rep1	rep2	ctl1
Unpaired Reads	0	0	0
Paired Reads	5186843	10670123	5625114
Unmapped Reads	0	0	0
Unpaired Duplicate Reads	0	0	0
Paired Duplicate Reads	713027	1749038	680113
Paired Optical Duplicate Reads	30440	97913	33270
% Duplicate Reads	13.7468	16.3919	12.0907

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	8947632	17842170	9890002
Total Reads (QC-failed)	0	0	0
Duplicate Reads	0	0	0
Duplicate Reads (QC-failed)	0	0	0
Mapped Reads	8947632	17842170	9890002
Mapped Reads (QC-failed)	0	0	0
% Mapped Reads	100.0	100.0	100.0
Paired Reads	8947632	17842170	9890002
Paired Reads (QC-failed)	0	0	0
Read1	4473816	8921085	4945001
Read1 (QC-failed)	0	0	0
Read2	4473816	8921085	4945001
Read2 (QC-failed)	0	0	0
Properly Paired Reads	8947632	17842170	9890002
Properly Paired Reads (QC-failed)	0	0	0
% Properly Paired Reads	100.0	100.0	100.0
With itself	8947632	17842170	9890002
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	5120897	10485239	5545972
Distinct Fragments	4420202	8793331	4879387
Positions with Two Read	549296	1259146	541345
NRF = Distinct/Total	0.863169	0.838639	0.879807
PBC1 = OneRead/Distinct	0.85962	0.833929	0.876868
PBC2 = OneRead/TwoRead	6.917391	5.8238	7.903605

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	63174	2482
N1	29472	1495
N2	65886	1846
Np	68454	2847
N optimal	68454	2847
N conservative	63174	2482
Optimal Set	pooled-pr1_vs_pooled-pr2	pooled-pr1_vs_pooled-pr2
Conservative Set	rep1_vs_rep2	rep1_vs_rep2
Rescue Ratio	1.0835786874347042	1.1470588235294117
Self Consistency Ratio	2.235545602605863	1.2347826086956522
Reproducibility Test	borderline	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	95102	109473

Top 300000 raw peaks from spp with FDR 0.01

Peak calling statistics

Peak region size

	rep1	rep2	idr_opt	overlap_opt
Min size	82.0	76.0	90.0	90.0
25 percentile	330.0	304.0	360.0	360.0
50 percentile (median)	330.0	304.0	360.0	360.0
75 percentile	330.0	304.0	360.0	360.0
Max size	428.0	433.0	515.0	515.0
Mean	329.6631195979054	303.7868515524376	342.66034422198805	359.26219066818595

Enrichment / Signal-to-noise ratio

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

	rep1	rep2
Number of Subsampled Reads	4823652	9885769
Estimated Fragment Length	200	150
Cross-correlation at Estimated Fragment Length	0.597163006977034	0.743011045406639
Phantom Peak	50	50
Cross-correlation at Phantom Peak	0.5957637	0.7419605
Argmin of Cross-correlation	1500	1500
Minimum of Cross-correlation	0.5896796	0.7359334
NSC (Normalized Strand Cross-correlation coeff.)	1.012691	1.009617
RSC (Relative Strand Cross-correlation coeff.)	1.229987	1.174312

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	0.38155623124512394	0.3948668667133155
Synthetic AUC	0.4978673493451548	0.49847871401122795
X-intercept	0.019741346443663307	0.019128546306332202
Synthetic X-intercept	0.0	0.0
Elbow Point	0.5899608566426936	0.5650496328189405
Synthetic Elbow Point	0.5005451640994383	0.49883418741834157
JS Distance	0.08103717860334284	0.057223214616995706
Synthetic JS Distance	0.18182842402576482	0.16929628889561477
% Genome Enriched	37.60257415979512	39.716235610674296
Diff. Enrichment	13.078998551631166	10.862986688711734
CHANCE Divergence	0.11146522661517881	0.09242071239889048

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	0.47731209777067274	0.48955373701741434	0.39540338717551193	0.504171241035004	0.38854883616134417	0.505675767653348	0.47342981482281954	0.4895083218973905	0.4917776915094551

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	0.33915136812134705	0.20057541481366242	0.33445780417964854	0.3699515957602076

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	0.03024852516640474	0.023876596623553584	0.029039797289231076	0.0409214670567554

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