QC Report

	general
Report generated at	2024-02-13 13:45:36
Title	pqn-47_PHX2564_L2larva_1_1
Description	mkudron
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	24629926	25429632	25020262
Total Reads (QC-failed)	0	0	0
Duplicate Reads	0	0	0
Duplicate Reads (QC-failed)	0	0	0
Mapped Reads	19467590	18269941	17250300
Mapped Reads (QC-failed)	0	0	0
% Mapped Reads	79.0	71.8	68.89999999999999
Paired Reads	24629926	25429632	25020262
Paired Reads (QC-failed)	0	0	0
Read1	12314963	12714816	12510131
Read1 (QC-failed)	0	0	0
Read2	12314963	12714816	12510131
Read2 (QC-failed)	0	0	0
Properly Paired Reads	19411092	18215570	17172004
Properly Paired Reads (QC-failed)	0	0	0
% Properly Paired Reads	78.8	71.6	68.60000000000001
With itself	19456090	18257144	17232634
With itself (QC-failed)	0	0	0
Singletons	11500	12797	17666
Singletons (QC-failed)	0	0	0
% Singleton	0.0	0.1	0.1
Diff. Chroms	8020	8401	19192
Diff. Chroms (QC-failed)	0	0	0

Marking duplicates (filtered BAM)

	rep1	rep2	ctl1
Unpaired Reads	0	0	0
Paired Reads	8901932	8350842	7846503
Unmapped Reads	0	0	0
Unpaired Duplicate Reads	0	0	0
Paired Duplicate Reads	939892	847833	816534
Paired Optical Duplicate Reads	619183	534352	624415
% Duplicate Reads	10.5583	10.152700000000001	10.4063

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	15924080	15006018	14059938
Total Reads (QC-failed)	0	0	0
Duplicate Reads	0	0	0
Duplicate Reads (QC-failed)	0	0	0
Mapped Reads	15924080	15006018	14059938
Mapped Reads (QC-failed)	0	0	0
% Mapped Reads	100.0	100.0	100.0
Paired Reads	15924080	15006018	14059938
Paired Reads (QC-failed)	0	0	0
Read1	7962040	7503009	7029969
Read1 (QC-failed)	0	0	0
Read2	7962040	7503009	7029969
Read2 (QC-failed)	0	0	0
Properly Paired Reads	15924080	15006018	14059938
Properly Paired Reads (QC-failed)	0	0	0
% Properly Paired Reads	100.0	100.0	100.0
With itself	15924080	15006018	14059938
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	8859851	8314070	7804999
Distinct Fragments	7914017	7447353	6981451
Positions with Two Read	707866	648752	603233
NRF = Distinct/Total	0.893245	0.895753	0.894485
PBC1 = OneRead/Distinct	0.897149	0.900436	0.899691
PBC2 = OneRead/TwoRead	10.030223	10.336566	10.412472

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	18881	1917
N1	22546	1652
N2	18841	1214
Np	19275	2018
N optimal	19275	2018
N conservative	18881	1917
Optimal Set	pooled-pr1_vs_pooled-pr2	pooled-pr1_vs_pooled-pr2
Conservative Set	rep1_vs_rep2	rep1_vs_rep2
Rescue Ratio	1.0208675387956145	1.0526864893062076
Self Consistency Ratio	1.196645613290165	1.3607907742998353
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	49409	35628

Top 300000 raw peaks from spp with FDR 0.01

Peak calling statistics

Peak region size

	rep1	rep2	idr_opt	overlap_opt
Min size	84.0	79.0	86.0	86.0
25 percentile	336.0	316.0	344.0	344.0
50 percentile (median)	336.0	316.0	344.0	344.0
75 percentile	336.0	316.0	344.0	344.0
Max size	1370.0	1311.0	1400.0	1400.0
Mean	335.0899633669979	315.34711462894353	320.9846382556987	341.5825680933852

Enrichment / Signal-to-noise ratio

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

	rep1	rep2
Number of Subsampled Reads	8316047	7787980
Estimated Fragment Length	180	180
Cross-correlation at Estimated Fragment Length	0.727037400487315	0.714148735118851
Phantom Peak	50	50
Cross-correlation at Phantom Peak	0.723565	0.7116995
Argmin of Cross-correlation	1500	1500
Minimum of Cross-correlation	0.716204	0.7056318
NSC (Normalized Strand Cross-correlation coeff.)	1.015126	1.01207
RSC (Relative Strand Cross-correlation coeff.)	1.471739	1.403656

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	0.4072556222850872	0.4145046088869995
Synthetic AUC	0.4984017998705972	0.4983511812242824
X-intercept	0.018528757198031955	0.018540733126415906
Synthetic X-intercept	0.0	0.0
Elbow Point	0.5421203381203781	0.5255915609624654
Synthetic Elbow Point	0.49999618540602053	0.5008717122932076
JS Distance	0.06221009797500596	0.049740251859109144
Synthetic JS Distance	0.1470478492447087	0.13565646773606896
% Genome Enriched	39.88203710541811	41.37363898563886
Diff. Enrichment	9.337358715998462	8.336838598509777
CHANCE Divergence	0.07947578407545867	0.07088595612574784

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	0.28965874323665797	0.20644997227112483	0.31855341093488604	0.29901386243654215	0.3131247770671838	0.29752040781510564	0.21756581566602215	0.2412673091907236	0.244530505175283

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	0.12633901127633027	0.1531474345770682	0.12063959939272365	0.12877356547657884

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	0.02835933465196263	0.027845376310593768	0.019586741799190163	0.029226806846845425

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