QC Report

	general
Report generated at	2021-08-29 14:34:55
Title	egl-43_RW12181_L4larva_1_4
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	34630488	32720606	37778924
Total Reads (QC-failed)	0	0	0
Duplicate Reads	0	0	0
Duplicate Reads (QC-failed)	0	0	0
Mapped Reads	33348157	31655104	37649598
Mapped Reads (QC-failed)	0	0	0
% Mapped Reads	96.3	96.7	99.7
Paired Reads	34630488	32720606	37778924
Paired Reads (QC-failed)	0	0	0
Read1	17315244	16360303	18889462
Read1 (QC-failed)	0	0	0
Read2	17315244	16360303	18889462
Read2 (QC-failed)	0	0	0
Properly Paired Reads	33219260	31534986	37269468
Properly Paired Reads (QC-failed)	0	0	0
% Properly Paired Reads	95.89999999999999	96.39999999999999	98.7
With itself	33327018	31634848	37631304
With itself (QC-failed)	0	0	0
Singletons	21139	20256	18294
Singletons (QC-failed)	0	0	0
% Singleton	0.1	0.1	0.0
Diff. Chroms	8643	8393	168622
Diff. Chroms (QC-failed)	0	0	0

Marking duplicates (filtered BAM)

	rep1	rep2	ctl1
Unpaired Reads	0	0	0
Paired Reads	15187353	14404395	16890023
Unmapped Reads	0	0	0
Unpaired Duplicate Reads	0	0	0
Paired Duplicate Reads	2804675	2639510	2397150
Paired Optical Duplicate Reads	93984	93541	108974
% Duplicate Reads	18.467200000000002	18.324299999999997	14.1927

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	24765356	23529770	28985746
Total Reads (QC-failed)	0	0	0
Duplicate Reads	0	0	0
Duplicate Reads (QC-failed)	0	0	0
Mapped Reads	24765356	23529770	28985746
Mapped Reads (QC-failed)	0	0	0
% Mapped Reads	100.0	100.0	100.0
Paired Reads	24765356	23529770	28985746
Paired Reads (QC-failed)	0	0	0
Read1	12382678	11764885	14492873
Read1 (QC-failed)	0	0	0
Read2	12382678	11764885	14492873
Read2 (QC-failed)	0	0	0
Properly Paired Reads	24765356	23529770	28985746
Properly Paired Reads (QC-failed)	0	0	0
% Properly Paired Reads	100.0	100.0	100.0
With itself	24765356	23529770	28985746
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	15085775	14308583	16667098
Distinct Fragments	12310351	11696109	14324702
Positions with Two Read	1953755	1842046	1837372
NRF = Distinct/Total	0.816024	0.817419	0.85946
PBC1 = OneRead/Distinct	0.810806	0.812372	0.855232
PBC2 = OneRead/TwoRead	5.108783	5.158171	6.667642

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	52236	3024
N1	52365	1532
N2	49005	1621
Np	52864	3146
N optimal	52864	3146
N conservative	52236	3024
Optimal Set	pooled-pr1_vs_pooled-pr2	pooled-pr1_vs_pooled-pr2
Conservative Set	rep1_vs_rep2	rep1_vs_rep2
Rescue Ratio	1.0120223600581975	1.0403439153439153
Self Consistency Ratio	1.0685644322007959	1.0580939947780679
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	81290	74196

Top 300000 raw peaks from spp with FDR 0.01

Peak calling statistics

Peak region size

	rep1	rep2	idr_opt	overlap_opt
Min size	78.0	81.0	82.0	82.0
25 percentile	310.0	324.0	330.0	330.0
50 percentile (median)	310.0	324.0	330.0	330.0
75 percentile	310.0	324.0	330.0	330.0
Max size	360.0	438.0	585.0	585.0
Mean	309.37216139746585	323.1716130249609	305.251430387794	328.4660260290557

Enrichment / Signal-to-noise ratio

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

	rep1	rep2
Number of Subsampled Reads	14206459	13478066
Estimated Fragment Length	160	175
Cross-correlation at Estimated Fragment Length	0.801168490576195	0.79260349352406
Phantom Peak	55	50
Cross-correlation at Phantom Peak	0.797133	0.7871554
Argmin of Cross-correlation	1500	1500
Minimum of Cross-correlation	0.7897909	0.7796502
NSC (Normalized Strand Cross-correlation coeff.)	1.014406	1.016614
RSC (Relative Strand Cross-correlation coeff.)	1.549633	1.725914

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	0.3859455516342832	0.37872572252691195
Synthetic AUC	0.4987181881748846	0.49868497008598117
X-intercept	0.01860396338356337	0.018534023671595085
Synthetic X-intercept	0.0	0.0
Elbow Point	0.5310722157500235	0.54136534307427
Synthetic Elbow Point	0.49871898616746124	0.49894023091285306
JS Distance	0.08888787091577587	0.09946160650680358
Synthetic JS Distance	0.17103711317041878	0.1804527639959636
% Genome Enriched	42.311727291075094	41.6868659215756
Diff. Enrichment	11.002222477510754	12.194445338433669
CHANCE Divergence	0.0936453881137522	0.10380579014423731

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	0.4119140060009636	0.39848944549819226	0.4512451991402829	0.43794474506595304	0.4524569725547252	0.43667017881349274	0.3767516622691905	0.4065058902007673	0.4072772646779

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	0.3020315341966392	0.29679843891604063	0.296327120919584	0.305016762975212

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	0.029635371486555394	0.017151782514250955	0.018802861226437827	0.030572194800775547

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