QC Report

	general
Report generated at	2021-08-30 17:50:16
Title	let-526_RW12306_youngadult_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	21201858	19651538	21179644
Total Reads (QC-failed)	0	0	0
Duplicate Reads	0	0	0
Duplicate Reads (QC-failed)	0	0	0
Mapped Reads	21149053	19580763	21140203
Mapped Reads (QC-failed)	0	0	0
% Mapped Reads	99.8	99.6	99.8
Paired Reads	21201858	19651538	21179644
Paired Reads (QC-failed)	0	0	0
Read1	10600929	9825769	10589822
Read1 (QC-failed)	0	0	0
Read2	10600929	9825769	10589822
Read2 (QC-failed)	0	0	0
Properly Paired Reads	21052402	19468444	21016156
Properly Paired Reads (QC-failed)	0	0	0
% Properly Paired Reads	99.3	99.1	99.2
With itself	21130664	19551074	21120358
With itself (QC-failed)	0	0	0
Singletons	18389	29689	19845
Singletons (QC-failed)	0	0	0
% Singleton	0.1	0.2	0.1
Diff. Chroms	7836	8279	32807
Diff. Chroms (QC-failed)	0	0	0

Marking duplicates (filtered BAM)

	rep1	rep2	ctl1
Unpaired Reads	0	0	0
Paired Reads	9637305	8894146	9643994
Unmapped Reads	0	0	0
Unpaired Duplicate Reads	0	0	0
Paired Duplicate Reads	1766820	1588399	1757190
Paired Optical Duplicate Reads	164653	143955	179659
% Duplicate Reads	18.333099999999998	17.8589	18.2206

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	15740970	14611494	15773608
Total Reads (QC-failed)	0	0	0
Duplicate Reads	0	0	0
Duplicate Reads (QC-failed)	0	0	0
Mapped Reads	15740970	14611494	15773608
Mapped Reads (QC-failed)	0	0	0
% Mapped Reads	100.0	100.0	100.0
Paired Reads	15740970	14611494	15773608
Paired Reads (QC-failed)	0	0	0
Read1	7870485	7305747	7886804
Read1 (QC-failed)	0	0	0
Read2	7870485	7305747	7886804
Read2 (QC-failed)	0	0	0
Properly Paired Reads	15740970	14611494	15773608
Properly Paired Reads (QC-failed)	0	0	0
% Properly Paired Reads	100.0	100.0	100.0
With itself	15740970	14611494	15773608
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	9582775	8843144	9554715
Distinct Fragments	7837086	7279170	7822740
Positions with Two Read	1237303	1130114	1241145
NRF = Distinct/Total	0.817831	0.823143	0.818731
PBC1 = OneRead/Distinct	0.812332	0.817251	0.812345
PBC2 = OneRead/TwoRead	5.145315	5.263988	5.120079

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	55709	1077
N1	32966	585
N2	35964	421
Np	58377	1200
N optimal	58377	1200
N conservative	55709	1077
Optimal Set	pooled-pr1_vs_pooled-pr2	pooled-pr1_vs_pooled-pr2
Conservative Set	rep1_vs_rep2	rep1_vs_rep2
Rescue Ratio	1.0478917230609057	1.1142061281337048
Self Consistency Ratio	1.0909421828550627	1.3895486935866983
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	100146	98862

Top 300000 raw peaks from spp with FDR 0.01

Peak calling statistics

Peak region size

	rep1	rep2	idr_opt	overlap_opt
Min size	66.0	59.0	68.0	68.0
25 percentile	264.0	230.0	270.0	270.0
50 percentile (median)	264.0	230.0	270.0	270.0
75 percentile	264.0	230.0	270.0	270.0
Max size	264.0	230.0	282.0	282.0
Mean	263.92722624967547	229.97104044020958	260.85833333333335	269.8148243315004

Enrichment / Signal-to-noise ratio

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

	rep1	rep2
Number of Subsampled Reads	9040791	8330944
Estimated Fragment Length	145	130
Cross-correlation at Estimated Fragment Length	0.715994153263462	0.701256236011763
Phantom Peak	50	50
Cross-correlation at Phantom Peak	0.7160539	0.7014918
Argmin of Cross-correlation	1500	1500
Minimum of Cross-correlation	0.7113281	0.6967617
NSC (Normalized Strand Cross-correlation coeff.)	1.00656	1.006451
RSC (Relative Strand Cross-correlation coeff.)	0.9873483	0.9501897

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	0.4147327862267485	0.41273735290809
Synthetic AUC	0.4983892077505797	0.49830598798237813
X-intercept	0.019136281871342694	0.019483669773858462
Synthetic X-intercept	0.0	0.0
Elbow Point	0.5186760927246192	0.5139823630762597
Synthetic Elbow Point	0.5023343369559564	0.5005289923518184
JS Distance	0.043211018202486816	0.044158323802020776
Synthetic JS Distance	0.13089422926513666	0.13258629584892523
% Genome Enriched	43.877887162819114	43.996478205402084
Diff. Enrichment	8.171774608071825	8.471532512347608
CHANCE Divergence	0.06946004693901975	0.07202089883692503

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	0.4355631196806804	0.40095886156473803	0.3685224775191773	0.3761710642086204	0.3716094207166929	0.39374897512177404	0.527374252054133	0.3741386038196301	0.5094655919157788

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	0.25233065098108676	0.16530029597921855	0.16304048032323046	0.26236990183070474

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	0.010002944077291385	0.00657926417495237	0.0045073419596928285	0.010923561263428234

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