QC Report

	general
Report generated at	2021-08-30 16:53:53
Title	lin-1_RW12212_L4larva_2_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	9837592	9813910	13748000
Total Reads (QC-failed)	0	0	0
Duplicate Reads	0	0	0
Duplicate Reads (QC-failed)	0	0	0
Mapped Reads	9631679	9643977	13559276
Mapped Reads (QC-failed)	0	0	0
% Mapped Reads	97.89999999999999	98.3	98.6
Paired Reads	9837592	9813910	13748000
Paired Reads (QC-failed)	0	0	0
Read1	4918796	4906955	6874000
Read1 (QC-failed)	0	0	0
Read2	4918796	4906955	6874000
Read2 (QC-failed)	0	0	0
Properly Paired Reads	9591308	9600650	13482946
Properly Paired Reads (QC-failed)	0	0	0
% Properly Paired Reads	97.5	97.8	98.1
With itself	9624928	9637492	13548526
With itself (QC-failed)	0	0	0
Singletons	6751	6485	10750
Singletons (QC-failed)	0	0	0
% Singleton	0.1	0.1	0.1
Diff. Chroms	2381	2514	31186
Diff. Chroms (QC-failed)	0	0	0

Marking duplicates (filtered BAM)

	rep1	rep2	ctl1
Unpaired Reads	0	0	0
Paired Reads	4397794	4406190	6208531
Unmapped Reads	0	0	0
Unpaired Duplicate Reads	0	0	0
Paired Duplicate Reads	594091	547201	899674
Paired Optical Duplicate Reads	25111	22523	41822
% Duplicate Reads	13.5088	12.418899999999999	14.490900000000002

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	7607406	7717978	10617714
Total Reads (QC-failed)	0	0	0
Duplicate Reads	0	0	0
Duplicate Reads (QC-failed)	0	0	0
Mapped Reads	7607406	7717978	10617714
Mapped Reads (QC-failed)	0	0	0
% Mapped Reads	100.0	100.0	100.0
Paired Reads	7607406	7717978	10617714
Paired Reads (QC-failed)	0	0	0
Read1	3803703	3858989	5308857
Read1 (QC-failed)	0	0	0
Read2	3803703	3858989	5308857
Read2 (QC-failed)	0	0	0
Properly Paired Reads	7607406	7717978	10617714
Properly Paired Reads (QC-failed)	0	0	0
% Properly Paired Reads	100.0	100.0	100.0
With itself	7607406	7717978	10617714
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	4317773	4321107	6112864
Distinct Fragments	3739800	3790586	5233887
Positions with Two Read	452905	424660	687237
NRF = Distinct/Total	0.866141	0.877226	0.856209
PBC1 = OneRead/Distinct	0.863133	0.874762	0.851416
PBC2 = OneRead/TwoRead	7.127199	7.808268	6.484249

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	31866	2267
N1	27364	1079
N2	25219	1348
Np	32438	2329
N optimal	32438	2329
N conservative	31866	2267
Optimal Set	pooled-pr1_vs_pooled-pr2	pooled-pr1_vs_pooled-pr2
Conservative Set	rep1_vs_rep2	rep1_vs_rep2
Rescue Ratio	1.0179501663214712	1.027348919276577
Self Consistency Ratio	1.0850549189103453	1.2493049119555144
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	72230	60873

Top 300000 raw peaks from spp with FDR 0.01

Peak calling statistics

Peak region size

	rep1	rep2	idr_opt	overlap_opt
Min size	65.0	70.0	74.0	74.0
25 percentile	260.0	280.0	296.0	296.0
50 percentile (median)	260.0	280.0	296.0	296.0
75 percentile	260.0	280.0	296.0	296.0
Max size	260.0	280.0	443.0	443.0
Mean	259.91849646961094	279.81558326351586	286.3975955345642	295.31056168691043

Enrichment / Signal-to-noise ratio

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

	rep1	rep2
Number of Subsampled Reads	4075371	4086032
Estimated Fragment Length	140	150
Cross-correlation at Estimated Fragment Length	0.555911205204519	0.561769370212943
Phantom Peak	50	55
Cross-correlation at Phantom Peak	0.5557078	0.5606536
Argmin of Cross-correlation	1500	1500
Minimum of Cross-correlation	0.5503711	0.555236
NSC (Normalized Strand Cross-correlation coeff.)	1.010066	1.011767
RSC (Relative Strand Cross-correlation coeff.)	1.038109	1.205955

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	0.38382783304910967	0.38162795148148426
Synthetic AUC	0.4976753844285149	0.4976957022334227
X-intercept	0.020165088691264987	0.02025689333099165
Synthetic X-intercept	0.0	0.0
Elbow Point	0.5758944965114237	0.5845480816821803
Synthetic Elbow Point	0.4988867136957764	0.49802234870836126
JS Distance	0.06890860897533412	0.07400589372601384
Synthetic JS Distance	0.180346459424574	0.18431068968052436
% Genome Enriched	39.230118308240066	39.3069548001852
Diff. Enrichment	12.54531432670375	12.752790360633448
CHANCE Divergence	0.10674422798886878	0.1084975263066461

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	0.3694016068026342	0.33885312966686354	0.40332160441506487	0.38581856910746076	0.39895580673775183	0.3865674109377899	0.47745107072031606	0.3547347708260306	0.3530696139345321

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	0.19703049528807892	0.1713032011174374	0.16988257805347462	0.19907468550217078

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	0.03730281733886733	0.025586513983873085	0.028846674608297665	0.037907891900131184

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