QC Report

	general
Report generated at	2021-08-31 20:08:51
Title	ref-1_RW12224_midembryonic_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	17968476	21195490	20022208
Total Reads (QC-failed)	0	0	0
Duplicate Reads	0	0	0
Duplicate Reads (QC-failed)	0	0	0
Mapped Reads	17917782	21129650	19950836
Mapped Reads (QC-failed)	0	0	0
% Mapped Reads	99.7	99.7	99.6
Paired Reads	17968476	21195490	20022208
Paired Reads (QC-failed)	0	0	0
Read1	8984238	10597745	10011104
Read1 (QC-failed)	0	0	0
Read2	8984238	10597745	10011104
Read2 (QC-failed)	0	0	0
Properly Paired Reads	17792906	21003618	19555260
Properly Paired Reads (QC-failed)	0	0	0
% Properly Paired Reads	99.0	99.1	97.7
With itself	17893348	21096426	19893556
With itself (QC-failed)	0	0	0
Singletons	24434	33224	57280
Singletons (QC-failed)	0	0	0
% Singleton	0.1	0.2	0.3
Diff. Chroms	5806	7568	11085
Diff. Chroms (QC-failed)	0	0	0

Marking duplicates (filtered BAM)

	rep1	rep2	ctl1
Unpaired Reads	0	0	0
Paired Reads	8028893	9438707	8965965
Unmapped Reads	0	0	0
Unpaired Duplicate Reads	0	0	0
Paired Duplicate Reads	1530881	1895633	1663348
Paired Optical Duplicate Reads	150305	190788	226584
% Duplicate Reads	19.0671	20.083599999999997	18.5518

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	12996024	15086148	14605234
Total Reads (QC-failed)	0	0	0
Duplicate Reads	0	0	0
Duplicate Reads (QC-failed)	0	0	0
Mapped Reads	12996024	15086148	14605234
Mapped Reads (QC-failed)	0	0	0
% Mapped Reads	100.0	100.0	100.0
Paired Reads	12996024	15086148	14605234
Paired Reads (QC-failed)	0	0	0
Read1	6498012	7543074	7302617
Read1 (QC-failed)	0	0	0
Read2	6498012	7543074	7302617
Read2 (QC-failed)	0	0	0
Properly Paired Reads	12996024	15086148	14605234
Properly Paired Reads (QC-failed)	0	0	0
% Properly Paired Reads	100.0	100.0	100.0
With itself	12996024	15086148	14605234
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	7993189	9390482	8915266
Distinct Fragments	6493002	7527767	7285919
Positions with Two Read	1071558	1302540	1186156
NRF = Distinct/Total	0.812317	0.801638	0.817241
PBC1 = OneRead/Distinct	0.804576	0.792859	0.808946
PBC2 = OneRead/TwoRead	4.875252	4.582167	4.968921

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	48678	675
N1	32460	344
N2	32530	405
Np	46316	653
N optimal	48678	675
N conservative	48678	675
Optimal Set	rep1_vs_rep2	rep1_vs_rep2
Conservative Set	rep1_vs_rep2	rep1_vs_rep2
Rescue Ratio	1.0509974954659298	1.0336906584992342
Self Consistency Ratio	1.0021565003080715	1.177325581395349
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	91453	90304

Top 300000 raw peaks from spp with FDR 0.01

Peak calling statistics

Peak region size

	rep1	rep2	idr_opt	overlap_opt
Min size	61.0	65.0	69.0	69.0
25 percentile	244.0	260.0	276.0	276.0
50 percentile (median)	244.0	260.0	276.0	276.0
75 percentile	244.0	260.0	276.0	276.0
Max size	244.0	263.0	292.0	292.0
Mean	243.928575333778	259.8834713855422	256.9303703703704	275.7355684292699

Enrichment / Signal-to-noise ratio

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

	rep1	rep2
Number of Subsampled Reads	7551818	8860127
Estimated Fragment Length	140	150
Cross-correlation at Estimated Fragment Length	0.675377787541199	0.705117353500099
Phantom Peak	50	50
Cross-correlation at Phantom Peak	0.6755208	0.7053657
Argmin of Cross-correlation	1500	1500
Minimum of Cross-correlation	0.6711855	0.7008572
NSC (Normalized Strand Cross-correlation coeff.)	1.006246	1.006079
RSC (Relative Strand Cross-correlation coeff.)	0.9670203	0.9449156

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	0.4134212772088991	0.4167718305310494
Synthetic AUC	0.4982196047914218	0.4983499810311976
X-intercept	0.019379891391151573	0.019152291966139594
Synthetic X-intercept	0.0	0.0
Elbow Point	0.5058337326305702	0.5053765372943619
Synthetic Elbow Point	0.49879444454233407	0.5010689751592452
JS Distance	0.033666314103424674	0.02875933235482715
Synthetic JS Distance	0.12890972618507732	0.12642705038206684
% Genome Enriched	44.48670340201246	43.99297236863121
Diff. Enrichment	8.27776449114912	7.771680370545853
CHANCE Divergence	0.07040127580694142	0.0660593420858131

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	0.38915871500391197	0.39138996912929663	0.3895933094614168	0.3683846930309844	0.3728817367527176	0.36044389860155157	0.5325290721814537	0.3792646808088776	0.36985223222762115

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	0.22381961765635508	0.15369600733270422	0.156953517889391	0.2145421657555548

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	0.0068523189730480966	0.004247222073458775	0.004951562188041639	0.006719067171869754

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