QC Report

	general
Report generated at	2022-07-25 10:02:09
Title	ceh-51_PHX1551_midembryonic_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	13456836	14719638	14478910
Total Reads (QC-failed)	0	0	0
Duplicate Reads	0	0	0
Duplicate Reads (QC-failed)	0	0	0
Mapped Reads	13401581	14675919	14447948
Mapped Reads (QC-failed)	0	0	0
% Mapped Reads	99.6	99.7	99.8
Paired Reads	13456836	14719638	14478910
Paired Reads (QC-failed)	0	0	0
Read1	6728418	7359819	7239455
Read1 (QC-failed)	0	0	0
Read2	6728418	7359819	7239455
Read2 (QC-failed)	0	0	0
Properly Paired Reads	13262684	14564266	14369296
Properly Paired Reads (QC-failed)	0	0	0
% Properly Paired Reads	98.6	98.9	99.2
With itself	13375694	14655138	14426304
With itself (QC-failed)	0	0	0
Singletons	25887	20781	21644
Singletons (QC-failed)	0	0	0
% Singleton	0.2	0.1	0.1
Diff. Chroms	26130	17441	10096
Diff. Chroms (QC-failed)	0	0	0

Marking duplicates (filtered BAM)

	rep1	rep2	ctl1
Unpaired Reads	0	0	0
Paired Reads	5559777	6210539	6584393
Unmapped Reads	0	0	0
Unpaired Duplicate Reads	0	0	0
Paired Duplicate Reads	852903	985661	933438
Paired Optical Duplicate Reads	47484	56288	61241
% Duplicate Reads	15.340599999999998	15.8708	14.1765

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	9413748	10449756	11301910
Total Reads (QC-failed)	0	0	0
Duplicate Reads	0	0	0
Duplicate Reads (QC-failed)	0	0	0
Mapped Reads	9413748	10449756	11301910
Mapped Reads (QC-failed)	0	0	0
% Mapped Reads	100.0	100.0	100.0
Paired Reads	9413748	10449756	11301910
Paired Reads (QC-failed)	0	0	0
Read1	4706874	5224878	5650955
Read1 (QC-failed)	0	0	0
Read2	4706874	5224878	5650955
Read2 (QC-failed)	0	0	0
Properly Paired Reads	9413748	10449756	11301910
Properly Paired Reads (QC-failed)	0	0	0
% Properly Paired Reads	100.0	100.0	100.0
With itself	9413748	10449756	11301910
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	5532055	6183552	6552186
Distinct Fragments	4690992	5209320	5631119
Positions with Two Read	629600	728061	717383
NRF = Distinct/Total	0.847966	0.842448	0.859426
PBC1 = OneRead/Distinct	0.845023	0.838399	0.855567
PBC2 = OneRead/TwoRead	6.296058	5.998794	6.715795

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	37496	1025
N1	25019	762
N2	22145	467
Np	39566	1158
N optimal	39566	1158
N conservative	37496	1025
Optimal Set	pooled-pr1_vs_pooled-pr2	pooled-pr1_vs_pooled-pr2
Conservative Set	rep1_vs_rep2	rep1_vs_rep2
Rescue Ratio	1.0552058886281204	1.1297560975609755
Self Consistency Ratio	1.1297809889365544	1.63169164882227
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	71853	49177

Top 300000 raw peaks from spp with FDR 0.01

Peak calling statistics

Peak region size

	rep1	rep2	idr_opt	overlap_opt
Min size	88.0	142.0	109.0	109.0
25 percentile	350.0	570.0	436.0	436.0
50 percentile (median)	350.0	570.0	436.0	436.0
75 percentile	350.0	570.0	436.0	436.0
Max size	398.0	570.0	452.0	452.0
Mean	349.814858113092	569.4525896252313	406.3117443868739	435.1303391801041

Enrichment / Signal-to-noise ratio

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

	rep1	rep2
Number of Subsampled Reads	5151533	5760294
Estimated Fragment Length	205	65
Cross-correlation at Estimated Fragment Length	0.604463348348879	0.627263152669198
Phantom Peak	50	50
Cross-correlation at Phantom Peak	0.6054065	0.62802
Argmin of Cross-correlation	1500	1500
Minimum of Cross-correlation	0.6004166	0.6233215
NSC (Normalized Strand Cross-correlation coeff.)	1.00674	1.006324
RSC (Relative Strand Cross-correlation coeff.)	0.8109838	0.8389264

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	0.41050651136942073	0.41527330912372185
Synthetic AUC	0.49791795914234843	0.4980240454806629
X-intercept	0.01876819503279346	0.018732278861138103
Synthetic X-intercept	0.0	0.0
Elbow Point	0.5255773025757882	0.5135453850712437
Synthetic Elbow Point	0.4998002144141095	0.5025733204468794
JS Distance	0.037232412314059396	0.0278996722591649
Synthetic JS Distance	0.13367204106547242	0.12648418632505262
% Genome Enriched	41.930733667619776	42.98986166287884
Diff. Enrichment	9.362395337554513	8.584332943337692
CHANCE Divergence	0.07958606519415841	0.0729535026814275

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	0.39351733231014896	0.34869589299501347	0.37532680925811907	0.3672847480840701	0.37802732769137226	0.3602038171991767	0.5366447430423152	0.4001393711804322	0.36377277644467965

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	0.219064320172312	0.16301976640972332	0.17517356386120403	0.23748700128637928

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	0.012857197803569803	0.013472742206398557	0.007815397794934158	0.014280108887132905

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