QC Report

	general
Report generated at	2023-11-22 17:45:24
Title	ceh-89_OH16505_L4larva_1_2
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	14956662	12751158	11223058
Total Reads (QC-failed)	0	0	0
Duplicate Reads	0	0	0
Duplicate Reads (QC-failed)	0	0	0
Mapped Reads	14788615	11936584	11030430
Mapped Reads (QC-failed)	0	0	0
% Mapped Reads	98.9	93.60000000000001	98.3
Paired Reads	14956662	12751158	11223058
Paired Reads (QC-failed)	0	0	0
Read1	7478331	6375579	5611529
Read1 (QC-failed)	0	0	0
Read2	7478331	6375579	5611529
Read2 (QC-failed)	0	0	0
Properly Paired Reads	14733022	11885718	10938824
Properly Paired Reads (QC-failed)	0	0	0
% Properly Paired Reads	98.5	93.2	97.5
With itself	14781066	11926488	11023960
With itself (QC-failed)	0	0	0
Singletons	7549	10096	6470
Singletons (QC-failed)	0	0	0
% Singleton	0.1	0.1	0.1
Diff. Chroms	13630	10529	48809
Diff. Chroms (QC-failed)	0	0	0

Marking duplicates (filtered BAM)

	rep1	rep2	ctl1
Unpaired Reads	0	0	0
Paired Reads	6698259	5402819	4956437
Unmapped Reads	0	0	0
Unpaired Duplicate Reads	0	0	0
Paired Duplicate Reads	1200818	1090825	612453
Paired Optical Duplicate Reads	835407	635314	500487
% Duplicate Reads	17.9273	20.189899999999998	12.3567

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	10994882	8623988	8687968
Total Reads (QC-failed)	0	0	0
Duplicate Reads	0	0	0
Duplicate Reads (QC-failed)	0	0	0
Mapped Reads	10994882	8623988	8687968
Mapped Reads (QC-failed)	0	0	0
% Mapped Reads	100.0	100.0	100.0
Paired Reads	10994882	8623988	8687968
Paired Reads (QC-failed)	0	0	0
Read1	5497441	4311994	4343984
Read1 (QC-failed)	0	0	0
Read2	5497441	4311994	4343984
Read2 (QC-failed)	0	0	0
Properly Paired Reads	10994882	8623988	8687968
Properly Paired Reads (QC-failed)	0	0	0
% Properly Paired Reads	100.0	100.0	100.0
With itself	10994882	8623988	8687968
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	6662772	5368649	4924524
Distinct Fragments	5478253	4294838	4322173
Positions with Two Read	766722	661510	439348
NRF = Distinct/Total	0.822218	0.799985	0.877683
PBC1 = OneRead/Distinct	0.826647	0.804335	0.881399
PBC2 = OneRead/TwoRead	5.906416	5.222128	8.670935

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	2630	417
N1	6880	204
N2	11241	303
Np	2329	420
N optimal	2630	420
N conservative	2630	417
Optimal Set	rep1_vs_rep2	pooled-pr1_vs_pooled-pr2
Conservative Set	rep1_vs_rep2	rep1_vs_rep2
Rescue Ratio	1.1292400171747532	1.0071942446043165
Self Consistency Ratio	1.6338662790697673	1.4852941176470589
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	7982	14837

Top 300000 raw peaks from spp with FDR 0.01

Peak calling statistics

Peak region size

	rep1	rep2	idr_opt	overlap_opt
Min size	80.0	80.0	84.0	84.0
25 percentile	316.0	320.0	330.0	330.0
50 percentile (median)	316.0	320.0	330.0	330.0
75 percentile	316.0	320.0	330.0	330.0
Max size	316.0	320.0	330.0	330.0
Mean	315.8408920070158	319.8647300667251	320.78333333333336	328.5281368821293

Enrichment / Signal-to-noise ratio

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

	rep1	rep2
Number of Subsampled Reads	6279256	5054466
Estimated Fragment Length	175	220
Cross-correlation at Estimated Fragment Length	0.633868796917205	0.578544986856468
Phantom Peak	50	50
Cross-correlation at Phantom Peak	0.633411	0.578092
Argmin of Cross-correlation	1500	1500
Minimum of Cross-correlation	0.6293514	0.5744231
NSC (Normalized Strand Cross-correlation coeff.)	1.007178	1.007176
RSC (Relative Strand Cross-correlation coeff.)	1.112774	1.123466

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	0.4107674412246122	0.40786846295373586
Synthetic AUC	0.4980752464124144	0.4978247183685452
X-intercept	0.018768660886781308	0.019131899538567164
Synthetic X-intercept	0.0	0.0
Elbow Point	0.5141543325190401	0.517912455493286
Synthetic Elbow Point	0.5018454865897787	0.5031656519886442
JS Distance	0.0148364523107382	0.01776429702045422
Synthetic JS Distance	0.13312347921757067	0.13745934743450883
% Genome Enriched	43.32898245277897	43.020828343791415
Diff. Enrichment	6.150768368775916	7.117355921136948
CHANCE Divergence	0.0523089702062652	0.06051512670438038

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	0.0550272390372175	0.09756147619871457	0.33350965776446573	0.3708200892672856	0.3448621540207806	0.3592370490311443	0.5844641409010815	0.06770226137364065	0.060964475626218595

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	0.02154986500241859	0.047949036651780345	0.07578210915877898	0.019855119076684843

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	0.005224103121127772	0.00305469399307787	0.005320160464045173	0.00525259609753263

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