QC Report

	general
Report generated at	2021-09-01 12:10:04
Title	Y75B8A.6_RW12275_L4larva_1_3
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	10911168	9645446	12631876
Total Reads (QC-failed)	0	0	0
Duplicate Reads	0	0	0
Duplicate Reads (QC-failed)	0	0	0
Mapped Reads	10871472	9603637	12599157
Mapped Reads (QC-failed)	0	0	0
% Mapped Reads	99.6	99.6	99.7
Paired Reads	10911168	9645446	12631876
Paired Reads (QC-failed)	0	0	0
Read1	5455584	4822723	6315938
Read1 (QC-failed)	0	0	0
Read2	5455584	4822723	6315938
Read2 (QC-failed)	0	0	0
Properly Paired Reads	10832056	9565250	12514536
Properly Paired Reads (QC-failed)	0	0	0
% Properly Paired Reads	99.3	99.2	99.1
With itself	10863424	9598020	12590086
With itself (QC-failed)	0	0	0
Singletons	8048	5617	9071
Singletons (QC-failed)	0	0	0
% Singleton	0.1	0.1	0.1
Diff. Chroms	3191	2279	39987
Diff. Chroms (QC-failed)	0	0	0

Marking duplicates (filtered BAM)

	rep1	rep2	ctl1
Unpaired Reads	0	0	0
Paired Reads	4973922	4387411	5770699
Unmapped Reads	0	0	0
Unpaired Duplicate Reads	0	0	0
Paired Duplicate Reads	620671	534549	766113
Paired Optical Duplicate Reads	28717	22928	35375
% Duplicate Reads	12.478499999999999	12.1837	13.275899999999998

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	8706502	7705724	10009172
Total Reads (QC-failed)	0	0	0
Duplicate Reads	0	0	0
Duplicate Reads (QC-failed)	0	0	0
Mapped Reads	8706502	7705724	10009172
Mapped Reads (QC-failed)	0	0	0
% Mapped Reads	100.0	100.0	100.0
Paired Reads	8706502	7705724	10009172
Paired Reads (QC-failed)	0	0	0
Read1	4353251	3852862	5004586
Read1 (QC-failed)	0	0	0
Read2	4353251	3852862	5004586
Read2 (QC-failed)	0	0	0
Properly Paired Reads	8706502	7705724	10009172
Properly Paired Reads (QC-failed)	0	0	0
% Properly Paired Reads	100.0	100.0	100.0
With itself	8706502	7705724	10009172
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	4938828	4356507	5683729
Distinct Fragments	4329278	3828517	4932720
Positions with Two Read	488775	427039	596820
NRF = Distinct/Total	0.87658	0.878804	0.867867
PBC1 = OneRead/Distinct	0.873898	0.875961	0.864209
PBC2 = OneRead/TwoRead	7.740467	7.853222	7.14269

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	71093	798
N1	44366	343
N2	36767	345
Np	75570	933
N optimal	75570	933
N conservative	71093	798
Optimal Set	pooled-pr1_vs_pooled-pr2	pooled-pr1_vs_pooled-pr2
Conservative Set	rep1_vs_rep2	rep1_vs_rep2
Rescue Ratio	1.0629738511527154	1.169172932330827
Self Consistency Ratio	1.2066799031740418	1.0058309037900874
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	114192	134410

Top 300000 raw peaks from spp with FDR 0.01

Peak calling statistics

Peak region size

	rep1	rep2	idr_opt	overlap_opt
Min size	88.0	57.0	64.0	64.0
25 percentile	350.0	224.0	256.0	256.0
50 percentile (median)	350.0	224.0	256.0	256.0
75 percentile	350.0	224.0	256.0	256.0
Max size	382.0	224.0	256.0	256.0
Mean	349.96523399187333	223.99473997470426	253.0042872454448	255.96076485377796

Enrichment / Signal-to-noise ratio

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

	rep1	rep2
Number of Subsampled Reads	4657822	4104244
Estimated Fragment Length	70	135
Cross-correlation at Estimated Fragment Length	0.592514299445733	0.563320813478906
Phantom Peak	50	50
Cross-correlation at Phantom Peak	0.5930082	0.5639294
Argmin of Cross-correlation	1500	1500
Minimum of Cross-correlation	0.5884855	0.5590938
NSC (Normalized Strand Cross-correlation coeff.)	1.006846	1.00756
RSC (Relative Strand Cross-correlation coeff.)	0.8908032	0.8741411

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	0.4019547182357788	0.39810192655059257
Synthetic AUC	0.4978182467350038	0.4976768474077653
X-intercept	0.020202100826921233	0.02028191947894384
Synthetic X-intercept	0.0	0.0
Elbow Point	0.5331167587241786	0.5358665112863574
Synthetic Elbow Point	0.5013912968500286	0.5025529125783742
JS Distance	0.051426720963468744	0.05562722842276033
Synthetic JS Distance	0.1460225859958177	0.15063170387387398
% Genome Enriched	43.66619041537626	43.176902078477696
Diff. Enrichment	10.203628125234193	10.79278381750211
CHANCE Divergence	0.08678721486953189	0.09179497865338256

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	0.5397321450107058	0.5245302063764546	0.4702378819328631	0.447477226020553	0.47743846551427094	0.44048735719057674	0.5290879494347689	0.5340649666821592	0.5324865173665677

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	0.3063808041639202	0.2595507357604696	0.17415469331629319	0.32302723591546934

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	0.008597919624065621	0.004796989652101384	0.004310821410162108	0.009633367222703368

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