QC Report

	general
Report generated at	2021-08-31 17:25:25
Title	php-3_RW12213_L4larva_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	24225544	26626118	24469960
Total Reads (QC-failed)	0	0	0
Duplicate Reads	0	0	0
Duplicate Reads (QC-failed)	0	0	0
Mapped Reads	23919077	26243996	24091108
Mapped Reads (QC-failed)	0	0	0
% Mapped Reads	98.7	98.6	98.5
Paired Reads	24225544	26626118	24469960
Paired Reads (QC-failed)	0	0	0
Read1	12112772	13313059	12234980
Read1 (QC-failed)	0	0	0
Read2	12112772	13313059	12234980
Read2 (QC-failed)	0	0	0
Properly Paired Reads	23815776	26158062	23993154
Properly Paired Reads (QC-failed)	0	0	0
% Properly Paired Reads	98.3	98.2	98.1
With itself	23902812	26225518	24064274
With itself (QC-failed)	0	0	0
Singletons	16265	18478	26834
Singletons (QC-failed)	0	0	0
% Singleton	0.1	0.1	0.1
Diff. Chroms	6313	7461	16202
Diff. Chroms (QC-failed)	0	0	0

Marking duplicates (filtered BAM)

	rep1	rep2	ctl1
Unpaired Reads	0	0	0
Paired Reads	10943151	12017766	11036161
Unmapped Reads	0	0	0
Unpaired Duplicate Reads	0	0	0
Paired Duplicate Reads	1123510	1334684	889419
Paired Optical Duplicate Reads	61831	74282	58470
% Duplicate Reads	10.2668	11.1059	8.059099999999999

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	19639282	21366164	20293484
Total Reads (QC-failed)	0	0	0
Duplicate Reads	0	0	0
Duplicate Reads (QC-failed)	0	0	0
Mapped Reads	19639282	21366164	20293484
Mapped Reads (QC-failed)	0	0	0
% Mapped Reads	100.0	100.0	100.0
Paired Reads	19639282	21366164	20293484
Paired Reads (QC-failed)	0	0	0
Read1	9819641	10683082	10146742
Read1 (QC-failed)	0	0	0
Read2	9819641	10683082	10146742
Read2 (QC-failed)	0	0	0
Properly Paired Reads	19639282	21366164	20293484
Properly Paired Reads (QC-failed)	0	0	0
% Properly Paired Reads	100.0	100.0	100.0
With itself	19639282	21366164	20293484
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	10856964	11929042	10901388
Distinct Fragments	9746702	10609485	10029780
Positions with Two Read	926052	1084830	766058
NRF = Distinct/Total	0.897737	0.889383	0.920046
PBC1 = OneRead/Distinct	0.89601	0.887275	0.918584
PBC2 = OneRead/TwoRead	9.430513	8.677422	12.026756

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	73133	3495
N1	32617	1728
N2	38357	2000
Np	73291	3559
N optimal	73291	3559
N conservative	73133	3495
Optimal Set	pooled-pr1_vs_pooled-pr2	pooled-pr1_vs_pooled-pr2
Conservative Set	rep1_vs_rep2	rep1_vs_rep2
Rescue Ratio	1.0021604474040446	1.0183118741058654
Self Consistency Ratio	1.1759818499555446	1.1574074074074074
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	118148	112727

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	75.0	81.0	81.0
25 percentile	284.0	300.0	324.0	324.0
50 percentile (median)	284.0	300.0	324.0	324.0
75 percentile	284.0	300.0	324.0	324.0
Max size	562.0	572.0	938.0	938.0
Mean	283.7926075769374	299.6930726445306	306.49592582186006	323.14195467383445

Enrichment / Signal-to-noise ratio

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

	rep1	rep2
Number of Subsampled Reads	10253464	11248661
Estimated Fragment Length	165	180
Cross-correlation at Estimated Fragment Length	0.77033226386475	0.783930038922975
Phantom Peak	50	50
Cross-correlation at Phantom Peak	0.7698502	0.7832704
Argmin of Cross-correlation	1500	1500
Minimum of Cross-correlation	0.7647707	0.7780885
NSC (Normalized Strand Cross-correlation coeff.)	1.007272	1.007508
RSC (Relative Strand Cross-correlation coeff.)	1.0949	1.127285

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	0.4152291003103257	0.41580918539531686
Synthetic AUC	0.4985613917111365	0.49862123654285034
X-intercept	0.01889538365714012	0.01870173841522331
Synthetic X-intercept	0.0	0.0
Elbow Point	0.5408252082185436	0.5424262750640826
Synthetic Elbow Point	0.5016643650652527	0.5000111286561354
JS Distance	0.04857549099788514	0.05025426823118301
Synthetic JS Distance	0.13547541285144066	0.1349226249241692
% Genome Enriched	41.80820736410895	41.55307476702681
Diff. Enrichment	8.06454149479926	8.05446666556745
CHANCE Divergence	0.06854983379639729	0.06847647024456595

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	0.49749664982660774	0.48784573590280406	0.3582274180667686	0.3790063579030845	0.34676688758447355	0.3815331568174802	0.4695949167337431	0.4939604610587354	0.4962099666571102

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	0.35095350505393846	0.17400330623084898	0.20508805417762402	0.3515386712291826

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	0.03180092224823015	0.01794643001714625	0.020753421156928308	0.032166702930142496

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