QC Report

	general
Report generated at	2021-08-31 14:51:05
Title	nhr-42_OP351_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	20761694	21987484	21707938
Total Reads (QC-failed)	0	0	0
Duplicate Reads	0	0	0
Duplicate Reads (QC-failed)	0	0	0
Mapped Reads	18420074	21545539	18848072
Mapped Reads (QC-failed)	0	0	0
% Mapped Reads	88.7	98.0	86.8
Paired Reads	20761694	21987484	21707938
Paired Reads (QC-failed)	0	0	0
Read1	10380847	10993742	10853969
Read1 (QC-failed)	0	0	0
Read2	10380847	10993742	10853969
Read2 (QC-failed)	0	0	0
Properly Paired Reads	18342504	21464474	18735068
Properly Paired Reads (QC-failed)	0	0	0
% Properly Paired Reads	88.3	97.6	86.3
With itself	18391968	21514744	18818212
With itself (QC-failed)	0	0	0
Singletons	28106	30795	29860
Singletons (QC-failed)	0	0	0
% Singleton	0.1	0.1	0.1
Diff. Chroms	8533	8028	16504
Diff. Chroms (QC-failed)	0	0	0

Marking duplicates (filtered BAM)

	rep1	rep2	ctl1
Unpaired Reads	0	0	0
Paired Reads	8426116	9857743	8621900
Unmapped Reads	0	0	0
Unpaired Duplicate Reads	0	0	0
Paired Duplicate Reads	1604103	1937494	1590014
Paired Optical Duplicate Reads	168563	200806	193651
% Duplicate Reads	19.0373	19.6545	18.4416

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	13644026	15840498	14063772
Total Reads (QC-failed)	0	0	0
Duplicate Reads	0	0	0
Duplicate Reads (QC-failed)	0	0	0
Mapped Reads	13644026	15840498	14063772
Mapped Reads (QC-failed)	0	0	0
% Mapped Reads	100.0	100.0	100.0
Paired Reads	13644026	15840498	14063772
Paired Reads (QC-failed)	0	0	0
Read1	6822013	7920249	7031886
Read1 (QC-failed)	0	0	0
Read2	6822013	7920249	7031886
Read2 (QC-failed)	0	0	0
Properly Paired Reads	13644026	15840498	14063772
Properly Paired Reads (QC-failed)	0	0	0
% Properly Paired Reads	100.0	100.0	100.0
With itself	13644026	15840498	14063772
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	8384947	9801136	8530240
Distinct Fragments	6801517	7896986	6974226
Positions with Two Read	1129084	1343065	1133748
NRF = Distinct/Total	0.811158	0.805722	0.817588
PBC1 = OneRead/Distinct	0.803279	0.797204	0.809274
PBC2 = OneRead/TwoRead	4.838894	4.687418	4.97823

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	80445	1627
N1	41909	875
N2	48557	702
Np	81215	1789
N optimal	81215	1789
N conservative	80445	1627
Optimal Set	pooled-pr1_vs_pooled-pr2	pooled-pr1_vs_pooled-pr2
Conservative Set	rep1_vs_rep2	rep1_vs_rep2
Rescue Ratio	1.009571757101125	1.0995697602950214
Self Consistency Ratio	1.1586294113436255	1.2464387464387465
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	125275	128405

Top 300000 raw peaks from spp with FDR 0.01

Peak calling statistics

Peak region size

	rep1	rep2	idr_opt	overlap_opt
Min size	69.0	69.0	76.0	76.0
25 percentile	276.0	276.0	300.0	300.0
50 percentile (median)	276.0	276.0	300.0	300.0
75 percentile	276.0	276.0	300.0	300.0
Max size	313.0	300.0	365.0	365.0
Mean	275.895877070445	275.9296211206729	290.1185019564002	299.77771347657455

Enrichment / Signal-to-noise ratio

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

	rep1	rep2
Number of Subsampled Reads	7914110	9265848
Estimated Fragment Length	155	150
Cross-correlation at Estimated Fragment Length	0.68568087745137	0.716257100097289
Phantom Peak	50	50
Cross-correlation at Phantom Peak	0.6854161	0.7163568
Argmin of Cross-correlation	1500	1500
Minimum of Cross-correlation	0.6806089	0.7118083
NSC (Normalized Strand Cross-correlation coeff.)	1.007452	1.00625
RSC (Relative Strand Cross-correlation coeff.)	1.055088	0.9780726

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	0.4074327427244985	0.4110976682765151
Synthetic AUC	0.49827430860816607	0.49839871115731643
X-intercept	0.018827996638071644	0.018668285077150668
Synthetic X-intercept	0.0	0.0
Elbow Point	0.5478525782436919	0.5422546880334117
Synthetic Elbow Point	0.4997163068609079	0.4987238621268691
JS Distance	0.04354749237045678	0.03571112175500601
Synthetic JS Distance	0.14146794442747268	0.13797294568254873
% Genome Enriched	43.68091227243598	43.89213081175397
Diff. Enrichment	8.711038583650204	8.174969328721804
CHANCE Divergence	0.07406810171208125	0.06950237452271753

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	0.5294724592286764	0.5308515552983246	0.42569789507907785	0.44575789905621666	0.43484429520206064	0.4519782713874616	0.5155542955348372	0.5307205867429861	0.5318409796055693

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	0.36710489204438235	0.2142357395097312	0.23606227531482912	0.3707387645125287

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	0.01652015816840048	0.011151840373215354	0.008344308367072803	0.017782786657841245

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