QC Report

	general
Report generated at	2022-05-31 23:13:26
Title	ZK337.2_RW12326_L4larva_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	13555512	16169108	12620906
Total Reads (QC-failed)	0	0	0
Duplicate Reads	0	0	0
Duplicate Reads (QC-failed)	0	0	0
Mapped Reads	13371012	16037753	12484485
Mapped Reads (QC-failed)	0	0	0
% Mapped Reads	98.6	99.2	98.9
Paired Reads	13555512	16169108	12620906
Paired Reads (QC-failed)	0	0	0
Read1	6777756	8084554	6310453
Read1 (QC-failed)	0	0	0
Read2	6777756	8084554	6310453
Read2 (QC-failed)	0	0	0
Properly Paired Reads	13307870	15955216	12422980
Properly Paired Reads (QC-failed)	0	0	0
% Properly Paired Reads	98.2	98.7	98.4
With itself	13345160	16008820	12466732
With itself (QC-failed)	0	0	0
Singletons	25852	28933	17753
Singletons (QC-failed)	0	0	0
% Singleton	0.2	0.2	0.1
Diff. Chroms	2946	3764	11662
Diff. Chroms (QC-failed)	0	0	0

Marking duplicates (filtered BAM)

	rep1	rep2	ctl1
Unpaired Reads	0	0	0
Paired Reads	6115009	7311553	5727363
Unmapped Reads	0	0	0
Unpaired Duplicate Reads	0	0	0
Paired Duplicate Reads	853895	1084174	616319
Paired Optical Duplicate Reads	84951	115504	79099
% Duplicate Reads	13.9639	14.828199999999999	10.761

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	10522228	12454758	10222088
Total Reads (QC-failed)	0	0	0
Duplicate Reads	0	0	0
Duplicate Reads (QC-failed)	0	0	0
Mapped Reads	10522228	12454758	10222088
Mapped Reads (QC-failed)	0	0	0
% Mapped Reads	100.0	100.0	100.0
Paired Reads	10522228	12454758	10222088
Paired Reads (QC-failed)	0	0	0
Read1	5261114	6227379	5111044
Read1 (QC-failed)	0	0	0
Read2	5261114	6227379	5111044
Read2 (QC-failed)	0	0	0
Properly Paired Reads	10522228	12454758	10222088
Properly Paired Reads (QC-failed)	0	0	0
% Properly Paired Reads	100.0	100.0	100.0
With itself	10522228	12454758	10222088
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	6013794	7226728	5563125
Distinct Fragments	5191937	6173867	4975738
Positions with Two Read	639313	811897	493306
NRF = Distinct/Total	0.863338	0.85431	0.894414
PBC1 = OneRead/Distinct	0.86037	0.850185	0.891823
PBC2 = OneRead/TwoRead	6.987169	6.465018	8.995388

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	98602	988
N1	45545	566
N2	66803	378
Np	99316	1043
N optimal	99316	1043
N conservative	98602	988
Optimal Set	pooled-pr1_vs_pooled-pr2	pooled-pr1_vs_pooled-pr2
Conservative Set	rep1_vs_rep2	rep1_vs_rep2
Rescue Ratio	1.0072412324293625	1.055668016194332
Self Consistency Ratio	1.4667471731254802	1.4973544973544974
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	151883	160875

Top 300000 raw peaks from spp with FDR 0.01

Peak calling statistics

Peak region size

	rep1	rep2	idr_opt	overlap_opt
Min size	56.0	56.0	60.0	60.0
25 percentile	224.0	224.0	240.0	240.0
50 percentile (median)	224.0	224.0	240.0	240.0
75 percentile	224.0	224.0	240.0	240.0
Max size	224.0	224.0	240.0	240.0
Mean	223.97180724636726	223.98907226107227	235.00479386385427	239.9475411816827

Enrichment / Signal-to-noise ratio

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

	rep1	rep2
Number of Subsampled Reads	5665535	6803663
Estimated Fragment Length	120	135
Cross-correlation at Estimated Fragment Length	0.63577628450776	0.672364246175633
Phantom Peak	50	50
Cross-correlation at Phantom Peak	0.6356843	0.6727146
Argmin of Cross-correlation	1500	1500
Minimum of Cross-correlation	0.6313669	0.6684039
NSC (Normalized Strand Cross-correlation coeff.)	1.006984	1.005925
RSC (Relative Strand Cross-correlation coeff.)	1.021299	0.9187349

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	0.4031682124352488	0.4139155287793446
Synthetic AUC	0.4980197317193603	0.4981826169112354
X-intercept	0.019854965138537635	0.019633400734156647
Synthetic X-intercept	0.0	0.0
Elbow Point	0.5407289269296564	0.5171193433709328
Synthetic Elbow Point	0.502939953509825	0.4998610675374095
JS Distance	0.042591777604940505	0.05282086216702356
Synthetic JS Distance	0.15539378953419278	0.13739013029691055
% Genome Enriched	46.78601868726084	47.97947235734546
Diff. Enrichment	8.143704151835829	6.847400473480636
CHANCE Divergence	0.0696389707022375	0.058600913631178815

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	0.5594159335836478	0.5763232011412827	0.44482822459273835	0.5280532101782772	0.45368490399561767	0.5247794497138282	0.5703500885625294	0.5669611700193254	0.5699441667365918

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	0.37527223979681235	0.19934941535195777	0.2660961377170074	0.3775030806912621

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	0.009327028357853375	0.006788961425279893	0.004116499092154179	0.009664017726258788

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