QC Report

	general
Report generated at	2023-08-04 13:06:48
Title	egl-38_OP171_L4larva_1_3
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	14804122	14353898	22053768
Total Reads (QC-failed)	0	0	0
Duplicate Reads	0	0	0
Duplicate Reads (QC-failed)	0	0	0
Mapped Reads	14580273	14287788	21991571
Mapped Reads (QC-failed)	0	0	0
% Mapped Reads	98.5	99.5	99.7
Paired Reads	14804122	14353898	22053768
Paired Reads (QC-failed)	0	0	0
Read1	7402061	7176949	11026884
Read1 (QC-failed)	0	0	0
Read2	7402061	7176949	11026884
Read2 (QC-failed)	0	0	0
Properly Paired Reads	14528752	14221852	21826196
Properly Paired Reads (QC-failed)	0	0	0
% Properly Paired Reads	98.1	99.1	99.0
With itself	14564772	14270814	21977720
With itself (QC-failed)	0	0	0
Singletons	15501	16974	13851
Singletons (QC-failed)	0	0	0
% Singleton	0.1	0.1	0.1
Diff. Chroms	6028	10792	67342
Diff. Chroms (QC-failed)	0	0	0

Marking duplicates (filtered BAM)

	rep1	rep2	ctl1
Unpaired Reads	0	0	0
Paired Reads	6647760	6513329	9970680
Unmapped Reads	0	0	0
Unpaired Duplicate Reads	0	0	0
Paired Duplicate Reads	739553	727000	1160926
Paired Optical Duplicate Reads	33057	46513	69778
% Duplicate Reads	11.1248	11.1617	11.6434

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	11816414	11572658	17619508
Total Reads (QC-failed)	0	0	0
Duplicate Reads	0	0	0
Duplicate Reads (QC-failed)	0	0	0
Mapped Reads	11816414	11572658	17619508
Mapped Reads (QC-failed)	0	0	0
% Mapped Reads	100.0	100.0	100.0
Paired Reads	11816414	11572658	17619508
Paired Reads (QC-failed)	0	0	0
Read1	5908207	5786329	8809754
Read1 (QC-failed)	0	0	0
Read2	5908207	5786329	8809754
Read2 (QC-failed)	0	0	0
Properly Paired Reads	11816414	11572658	17619508
Properly Paired Reads (QC-failed)	0	0	0
% Properly Paired Reads	100.0	100.0	100.0
With itself	11816414	11572658	17619508
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	6612058	6479969	9879555
Distinct Fragments	5883804	5761814	8737989
Positions with Two Read	595943	593238	940890
NRF = Distinct/Total	0.88986	0.889173	0.884452
PBC1 = OneRead/Distinct	0.888058	0.886743	0.881415
PBC2 = OneRead/TwoRead	8.767887	8.612479	8.185647

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	62828	1049
N1	37081	725
N2	33986	435
Np	62945	1153
N optimal	62945	1153
N conservative	62828	1049
Optimal Set	pooled-pr1_vs_pooled-pr2	pooled-pr1_vs_pooled-pr2
Conservative Set	rep1_vs_rep2	rep1_vs_rep2
Rescue Ratio	1.0018622270325332	1.0991420400381315
Self Consistency Ratio	1.0910669099040782	1.6666666666666667
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	114042	103529

Top 300000 raw peaks from spp with FDR 0.01

Peak calling statistics

Peak region size

	rep1	rep2	idr_opt	overlap_opt
Min size	72.0	76.0	78.0	78.0
25 percentile	290.0	304.0	310.0	310.0
50 percentile (median)	290.0	304.0	310.0	310.0
75 percentile	290.0	304.0	310.0	310.0
Max size	290.0	304.0	310.0	310.0
Mean	289.91831079777626	303.9580890378541	295.3694709453599	309.7283819207244

Enrichment / Signal-to-noise ratio

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

	rep1	rep2
Number of Subsampled Reads	6228437	6108315
Estimated Fragment Length	170	215
Cross-correlation at Estimated Fragment Length	0.666730065108939	0.660963825507826
Phantom Peak	50	50
Cross-correlation at Phantom Peak	0.6651622	0.6601583
Argmin of Cross-correlation	1500	1500
Minimum of Cross-correlation	0.6611806	0.6565452
NSC (Normalized Strand Cross-correlation coeff.)	1.008393	1.00673
RSC (Relative Strand Cross-correlation coeff.)	1.393785	1.222931

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	0.4077495884947235	0.4127601648267635
Synthetic AUC	0.49814821621829053	0.4981285841873942
X-intercept	0.01887383897838084	0.01881394104446024
Synthetic X-intercept	0.0	0.0
Elbow Point	0.5221941811153794	0.5089307819475614
Synthetic Elbow Point	0.5009203837272109	0.5017633898790522
JS Distance	0.05189179691713841	0.04435185955138231
Synthetic JS Distance	0.13895838680144493	0.13195990752232525
% Genome Enriched	43.423206855518195	44.36739795388658
Diff. Enrichment	9.94958742431782	9.107930219434268
CHANCE Divergence	0.08458567107135896	0.07744420564178907

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	0.514573541516064	0.4860252502061324	0.4282354311154922	0.41506308834788197	0.427528085513606	0.42162231384048743	0.5057941589131881	0.4971081371491546	0.49634684032728454

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	0.3108854425690767	0.2042387817488453	0.18857517434629106	0.3117733786103185

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	0.010724324590560925	0.008718719570929049	0.005102976343031998	0.011559757479903435

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