QC Report

	general
Report generated at	2021-08-30 11:45:27
Title	F26F4.8_OP776_L4larva_1_8
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	22006556	19366106	22060484
Total Reads (QC-failed)	0	0	0
Duplicate Reads	0	0	0
Duplicate Reads (QC-failed)	0	0	0
Mapped Reads	21945032	19305053	22000277
Mapped Reads (QC-failed)	0	0	0
% Mapped Reads	99.7	99.7	99.7
Paired Reads	22006556	19366106	22060484
Paired Reads (QC-failed)	0	0	0
Read1	11003278	9683053	11030242
Read1 (QC-failed)	0	0	0
Read2	11003278	9683053	11030242
Read2 (QC-failed)	0	0	0
Properly Paired Reads	21869674	19237850	21911022
Properly Paired Reads (QC-failed)	0	0	0
% Properly Paired Reads	99.4	99.3	99.3
With itself	21921666	19284640	21967150
With itself (QC-failed)	0	0	0
Singletons	23366	20413	33127
Singletons (QC-failed)	0	0	0
% Singleton	0.1	0.1	0.2
Diff. Chroms	6557	5888	18303
Diff. Chroms (QC-failed)	0	0	0

Marking duplicates (filtered BAM)

	rep1	rep2	ctl1
Unpaired Reads	0	0	0
Paired Reads	9983059	8783598	10056979
Unmapped Reads	0	0	0
Unpaired Duplicate Reads	0	0	0
Paired Duplicate Reads	2001178	1871942	1822307
Paired Optical Duplicate Reads	187910	186810	217503
% Duplicate Reads	20.0457	21.3118	18.1198

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	15963762	13823312	16469344
Total Reads (QC-failed)	0	0	0
Duplicate Reads	0	0	0
Duplicate Reads (QC-failed)	0	0	0
Mapped Reads	15963762	13823312	16469344
Mapped Reads (QC-failed)	0	0	0
% Mapped Reads	100.0	100.0	100.0
Paired Reads	15963762	13823312	16469344
Paired Reads (QC-failed)	0	0	0
Read1	7981881	6911656	8234672
Read1 (QC-failed)	0	0	0
Read2	7981881	6911656	8234672
Read2 (QC-failed)	0	0	0
Properly Paired Reads	15963762	13823312	16469344
Properly Paired Reads (QC-failed)	0	0	0
% Properly Paired Reads	100.0	100.0	100.0
With itself	15963762	13823312	16469344
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	9921886	8725576	9964247
Distinct Fragments	7954479	6885346	8180004
Positions with Two Read	1368101	1245244	1308163
NRF = Distinct/Total	0.80171	0.789099	0.820935
PBC1 = OneRead/Distinct	0.793493	0.77976	0.812974
PBC2 = OneRead/TwoRead	4.613567	4.311537	5.083563

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	82832	978
N1	42902	376
N2	39799	366
Np	82503	960
N optimal	82832	978
N conservative	82832	978
Optimal Set	rep1_vs_rep2	rep1_vs_rep2
Conservative Set	rep1_vs_rep2	rep1_vs_rep2
Rescue Ratio	1.0039877337793777	1.01875
Self Consistency Ratio	1.0779667830850022	1.0273224043715847
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	133622	137317

Top 300000 raw peaks from spp with FDR 0.01

Peak calling statistics

Peak region size

	rep1	rep2	idr_opt	overlap_opt
Min size	71.0	61.0	66.0	66.0
25 percentile	256.0	244.0	264.0	264.0
50 percentile (median)	256.0	244.0	264.0	264.0
75 percentile	256.0	244.0	264.0	264.0
Max size	256.0	244.0	264.0	264.0
Mean	255.98526440256845	243.98030105522258	258.51431492842534	263.9350854742129

Enrichment / Signal-to-noise ratio

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

	rep1	rep2
Number of Subsampled Reads	9350063	8223374
Estimated Fragment Length	170	145
Cross-correlation at Estimated Fragment Length	0.717100429275102	0.684695582564355
Phantom Peak	50	50
Cross-correlation at Phantom Peak	0.7172853	0.6848067
Argmin of Cross-correlation	1500	1500
Minimum of Cross-correlation	0.7130284	0.6807794
NSC (Normalized Strand Cross-correlation coeff.)	1.005711	1.005752
RSC (Relative Strand Cross-correlation coeff.)	0.9565799	0.9723989

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	0.41663230808900825	0.41567786776646026
Synthetic AUC	0.4984015087684163	0.4982796707047847
X-intercept	0.018834324228279836	0.0189041992916665
Synthetic X-intercept	0.0	0.0
Elbow Point	0.5056099693747579	0.5067000203635899
Synthetic Elbow Point	0.5014445259454835	0.5017903460830896
JS Distance	0.033025569303678066	0.033033663918259154
Synthetic JS Distance	0.12807825913121457	0.12956378464270618
% Genome Enriched	44.01390314118356	44.383442404979895
Diff. Enrichment	8.331423903680967	8.434428332993587
CHANCE Divergence	0.07081445922108928	0.07170669381889072

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	0.5306913871554838	0.5356793654082321	0.4103926868375153	0.41573625770727013	0.4217599111988654	0.41555988897595597	0.5181352152950639	0.5328876859212365	0.5325104797141526

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	0.3459712088538807	0.19934348808257102	0.18315060818999093	0.3445877228491795

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	0.007954524167093418	0.00381301099327339	0.0037560463078602294	0.00788795166655174

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