QC Report

	general
Report generated at	2022-05-06 19:46:25
Title	ceh-44_RW12327_L4larva_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	10166162	13956596	12480094
Total Reads (QC-failed)	0	0	0
Duplicate Reads	0	0	0
Duplicate Reads (QC-failed)	0	0	0
Mapped Reads	10106277	13898764	12437785
Mapped Reads (QC-failed)	0	0	0
% Mapped Reads	99.4	99.6	99.7
Paired Reads	10166162	13956596	12480094
Paired Reads (QC-failed)	0	0	0
Read1	5083081	6978298	6240047
Read1 (QC-failed)	0	0	0
Read2	5083081	6978298	6240047
Read2 (QC-failed)	0	0	0
Properly Paired Reads	10079090	13862276	12373576
Properly Paired Reads (QC-failed)	0	0	0
% Properly Paired Reads	99.1	99.3	99.1
With itself	10101740	13892632	12432386
With itself (QC-failed)	0	0	0
Singletons	4537	6132	5399
Singletons (QC-failed)	0	0	0
% Singleton	0.0	0.0	0.0
Diff. Chroms	2729	3735	11449
Diff. Chroms (QC-failed)	0	0	0

Marking duplicates (filtered BAM)

	rep1	rep2	ctl1
Unpaired Reads	0	0	0
Paired Reads	4628451	6364199	5698960
Unmapped Reads	0	0	0
Unpaired Duplicate Reads	0	0	0
Paired Duplicate Reads	514732	987497	655491
Paired Optical Duplicate Reads	18094	33244	25941
% Duplicate Reads	11.121	15.516399999999999	11.5019

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	8227438	10753404	10086938
Total Reads (QC-failed)	0	0	0
Duplicate Reads	0	0	0
Duplicate Reads (QC-failed)	0	0	0
Mapped Reads	8227438	10753404	10086938
Mapped Reads (QC-failed)	0	0	0
% Mapped Reads	100.0	100.0	100.0
Paired Reads	8227438	10753404	10086938
Paired Reads (QC-failed)	0	0	0
Read1	4113719	5376702	5043469
Read1 (QC-failed)	0	0	0
Read2	4113719	5376702	5043469
Read2 (QC-failed)	0	0	0
Properly Paired Reads	8227438	10753404	10086938
Properly Paired Reads (QC-failed)	0	0	0
% Properly Paired Reads	100.0	100.0	100.0
With itself	8227438	10753404	10086938
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	4585356	6302413	5580859
Distinct Fragments	4076552	5327292	4943155
Positions with Two Read	418112	735181	527667
NRF = Distinct/Total	0.889037	0.845278	0.885734
PBC1 = OneRead/Distinct	0.886872	0.840974	0.882676
PBC2 = OneRead/TwoRead	8.646918	6.093895	8.268855

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	85221	1003
N1	41983	387
N2	46136	525
Np	85848	1027
N optimal	85848	1027
N conservative	85221	1003
Optimal Set	pooled-pr1_vs_pooled-pr2	pooled-pr1_vs_pooled-pr2
Conservative Set	rep1_vs_rep2	rep1_vs_rep2
Rescue Ratio	1.0073573415003345	1.0239282153539382
Self Consistency Ratio	1.0989209918300265	1.3565891472868217
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	136434	145686

Top 300000 raw peaks from spp with FDR 0.01

Peak calling statistics

Peak region size

	rep1	rep2	idr_opt	overlap_opt
Min size	61.0	70.0	66.0	66.0
25 percentile	244.0	244.0	256.0	256.0
50 percentile (median)	244.0	244.0	256.0	256.0
75 percentile	244.0	244.0	256.0	256.0
Max size	244.0	244.0	256.0	256.0
Mean	243.99005379890644	243.99047266037917	253.27848101265823	255.96744245643464

Enrichment / Signal-to-noise ratio

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

	rep1	rep2
Number of Subsampled Reads	4321335	5943493
Estimated Fragment Length	150	150
Cross-correlation at Estimated Fragment Length	0.581690766809733	0.636397402222827
Phantom Peak	50	50
Cross-correlation at Phantom Peak	0.5820818	0.6362281
Argmin of Cross-correlation	1500	1500
Minimum of Cross-correlation	0.5776358	0.6322923
NSC (Normalized Strand Cross-correlation coeff.)	1.00702	1.006492
RSC (Relative Strand Cross-correlation coeff.)	0.91204	1.043017

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	0.40378739023522886	0.41158530625484485
Synthetic AUC	0.4977779054952787	0.4980555124205485
X-intercept	0.019987267222677914	0.019604086463141802
Synthetic X-intercept	0.0	0.0
Elbow Point	0.5246602763292082	0.5192737926313542
Synthetic Elbow Point	0.5011194363145958	0.498695478766013
JS Distance	0.04863356909285225	0.042624469448685316
Synthetic JS Distance	0.14519856300162756	0.13746370823118612
% Genome Enriched	46.691174269411995	45.91782768441073
Diff. Enrichment	9.611177721218562	8.208064860662267
CHANCE Divergence	0.08208420382055888	0.06993506385252908

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	0.5524204497195846	0.564809989469381	0.46769444687533424	0.4641717171604452	0.4721030950590196	0.4487641308742794	0.5607193822065428	0.5601045980884728	0.5594559566383784

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	0.3515957300524392	0.2018895797209289	0.2101107705057859	0.3539286613312518

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	0.009960358976698716	0.004906752259938027	0.006044318617620988	0.010160297419893174

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