QC Report

	general
Report generated at	2021-08-31 09:57:43
Title	nfya-2_RW12238_youngadult_1_4
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	10868264	14387350	11289296
Total Reads (QC-failed)	0	0	0
Duplicate Reads	0	0	0
Duplicate Reads (QC-failed)	0	0	0
Mapped Reads	10824073	14318913	11238554
Mapped Reads (QC-failed)	0	0	0
% Mapped Reads	99.6	99.5	99.6
Paired Reads	10868264	14387350	11289296
Paired Reads (QC-failed)	0	0	0
Read1	5434132	7193675	5644648
Read1 (QC-failed)	0	0	0
Read2	5434132	7193675	5644648
Read2 (QC-failed)	0	0	0
Properly Paired Reads	10786430	14280488	11184972
Properly Paired Reads (QC-failed)	0	0	0
% Properly Paired Reads	99.2	99.3	99.1
With itself	10816670	14308664	11226760
With itself (QC-failed)	0	0	0
Singletons	7403	10249	11794
Singletons (QC-failed)	0	0	0
% Singleton	0.1	0.1	0.1
Diff. Chroms	2600	2751	11885
Diff. Chroms (QC-failed)	0	0	0

Marking duplicates (filtered BAM)

	rep1	rep2	ctl1
Unpaired Reads	0	0	0
Paired Reads	4922879	6527459	5137479
Unmapped Reads	0	0	0
Unpaired Duplicate Reads	0	0	0
Paired Duplicate Reads	633155	1141932	711430
Paired Optical Duplicate Reads	34507	118842	82663
% Duplicate Reads	12.861500000000001	17.4943	13.8478

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	8579448	10771054	8852098
Total Reads (QC-failed)	0	0	0
Duplicate Reads	0	0	0
Duplicate Reads (QC-failed)	0	0	0
Mapped Reads	8579448	10771054	8852098
Mapped Reads (QC-failed)	0	0	0
% Mapped Reads	100.0	100.0	100.0
Paired Reads	8579448	10771054	8852098
Paired Reads (QC-failed)	0	0	0
Read1	4289724	5385527	4426049
Read1 (QC-failed)	0	0	0
Read2	4289724	5385527	4426049
Read2 (QC-failed)	0	0	0
Properly Paired Reads	8579448	10771054	8852098
Properly Paired Reads (QC-failed)	0	0	0
% Properly Paired Reads	100.0	100.0	100.0
With itself	8579448	10771054	8852098
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	4883434	6479981	5072076
Distinct Fragments	4258317	5355701	4374915
Positions with Two Read	509733	831057	549564
NRF = Distinct/Total	0.871992	0.826499	0.862549
PBC1 = OneRead/Distinct	0.867414	0.819309	0.858419
PBC2 = OneRead/TwoRead	7.246386	5.279992	6.833614

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	53004	281
N1	34383	92
N2	38482	90
Np	91644	435
N optimal	91644	435
N conservative	53004	281
Optimal Set	pooled-pr1_vs_pooled-pr2	pooled-pr1_vs_pooled-pr2
Conservative Set	rep1_vs_rep2	rep1_vs_rep2
Rescue Ratio	1.7290015847860538	1.5480427046263345
Self Consistency Ratio	1.1192158915743244	1.0222222222222221
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	74902	152501

Top 300000 raw peaks from spp with FDR 0.01

Peak calling statistics

Peak region size

	rep1	rep2	idr_opt	overlap_opt
Min size	58.0	92.0	69.0	69.0
25 percentile	230.0	236.0	240.0	240.0
50 percentile (median)	230.0	236.0	240.0	240.0
75 percentile	230.0	236.0	240.0	240.0
Max size	294.0	292.0	318.0	318.0
Mean	229.99627513284025	235.99758034373545	237.8919540229885	239.9899938893981

Enrichment / Signal-to-noise ratio

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

	rep1	rep2
Number of Subsampled Reads	4603751	6083117
Estimated Fragment Length	135	170
Cross-correlation at Estimated Fragment Length	0.587974077004305	0.633387727444394
Phantom Peak	50	55
Cross-correlation at Phantom Peak	0.5885162	0.6335737
Argmin of Cross-correlation	1500	1500
Minimum of Cross-correlation	0.5842153	0.6297289
NSC (Normalized Strand Cross-correlation coeff.)	1.006434	1.00581
RSC (Relative Strand Cross-correlation coeff.)	0.8739482	0.9516253

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	0.40536582369124924	0.4115249347727375
Synthetic AUC	0.49780091736854093	0.4980447075911146
X-intercept	0.02017817771880643	0.01981503835908896
Synthetic X-intercept	0.0	0.0
Elbow Point	0.5127667428195177	0.5022376966589184
Synthetic Elbow Point	0.5011278775632279	0.4992652843944219
JS Distance	0.035001953896449785	0.030804317712815117
Synthetic JS Distance	0.14169163664155465	0.1342866284713316
% Genome Enriched	44.65416961800533	46.06402825304029
Diff. Enrichment	9.430245904639445	8.481896221468183
CHANCE Divergence	0.08026935548411024	0.0722824069103933

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	0.32690110132959604	0.5726179629217345	0.4243345259508537	0.40864776861247404	0.41922930239800976	0.40052225167978023	0.5740922380204917	0.5590047680411838	0.5612478230536678

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	0.21948991297486753	0.15956189722229216	0.17224535314742642	0.3549037125755187

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	0.0028692795670107163	0.0013405291342753053	0.0012389688140083599	0.004014056069449774

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