QC Report

	general
Report generated at	2021-09-01 19:55:30
Title	ztf-23_RW12279_midembryonic_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	27094470	32463040	20674562
Total Reads (QC-failed)	0	0	0
Duplicate Reads	0	0	0
Duplicate Reads (QC-failed)	0	0	0
Mapped Reads	27012836	32379765	20644378
Mapped Reads (QC-failed)	0	0	0
% Mapped Reads	99.7	99.7	99.9
Paired Reads	27094470	32463040	20674562
Paired Reads (QC-failed)	0	0	0
Read1	13547235	16231520	10337281
Read1 (QC-failed)	0	0	0
Read2	13547235	16231520	10337281
Read2 (QC-failed)	0	0	0
Properly Paired Reads	26731674	32108542	20535814
Properly Paired Reads (QC-failed)	0	0	0
% Properly Paired Reads	98.7	98.9	99.3
With itself	26987594	32351300	20623624
With itself (QC-failed)	0	0	0
Singletons	25242	28465	20754
Singletons (QC-failed)	0	0	0
% Singleton	0.1	0.1	0.1
Diff. Chroms	17304	18017	13373
Diff. Chroms (QC-failed)	0	0	0

Marking duplicates (filtered BAM)

	rep1	rep2	ctl1
Unpaired Reads	0	0	0
Paired Reads	12019115	14389040	9433874
Unmapped Reads	0	0	0
Unpaired Duplicate Reads	0	0	0
Paired Duplicate Reads	1206489	1548577	776563
Paired Optical Duplicate Reads	73274	87656	53276
% Duplicate Reads	10.0381	10.7622	8.2316

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	21625252	25680926	17314622
Total Reads (QC-failed)	0	0	0
Duplicate Reads	0	0	0
Duplicate Reads (QC-failed)	0	0	0
Mapped Reads	21625252	25680926	17314622
Mapped Reads (QC-failed)	0	0	0
% Mapped Reads	100.0	100.0	100.0
Paired Reads	21625252	25680926	17314622
Paired Reads (QC-failed)	0	0	0
Read1	10812626	12840463	8657311
Read1 (QC-failed)	0	0	0
Read2	10812626	12840463	8657311
Read2 (QC-failed)	0	0	0
Properly Paired Reads	21625252	25680926	17314622
Properly Paired Reads (QC-failed)	0	0	0
% Properly Paired Reads	100.0	100.0	100.0
With itself	21625252	25680926	17314622
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	11941690	14294529	9386070
Distinct Fragments	10747539	12763578	8615808
Positions with Two Read	1008296	1265980	675374
NRF = Distinct/Total	0.900002	0.8929	0.917936
PBC1 = OneRead/Distinct	0.898011	0.891085	0.916341
PBC2 = OneRead/TwoRead	9.572004	8.983898	11.689841

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	80329	2234
N1	34972	959
N2	37194	1233
Np	80911	2413
N optimal	80911	2413
N conservative	80329	2234
Optimal Set	pooled-pr1_vs_pooled-pr2	pooled-pr1_vs_pooled-pr2
Conservative Set	rep1_vs_rep2	rep1_vs_rep2
Rescue Ratio	1.0072452040981463	1.0801253357206804
Self Consistency Ratio	1.0635365435205306	1.2857142857142858
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	119756	121116

Top 300000 raw peaks from spp with FDR 0.01

Peak calling statistics

Peak region size

	rep1	rep2	idr_opt	overlap_opt
Min size	81.0	76.0	86.0	86.0
25 percentile	324.0	304.0	344.0	344.0
50 percentile (median)	324.0	304.0	344.0	344.0
75 percentile	324.0	304.0	344.0	344.0
Max size	601.0	551.0	667.0	667.0
Mean	323.8239420154314	303.7560190230853	325.89888106092	343.4625452657859

Enrichment / Signal-to-noise ratio

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

	rep1	rep2
Number of Subsampled Reads	11300401	13504451
Estimated Fragment Length	160	155
Cross-correlation at Estimated Fragment Length	0.785905698251482	0.81235894444662
Phantom Peak	50	50
Cross-correlation at Phantom Peak	0.7862457	0.8122748
Argmin of Cross-correlation	1500	1500
Minimum of Cross-correlation	0.7812643	0.8070069
NSC (Normalized Strand Cross-correlation coeff.)	1.005941	1.006632
RSC (Relative Strand Cross-correlation coeff.)	0.9317481	1.015968

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	0.4208139023287647	0.4185042321062747
Synthetic AUC	0.4986295201380099	0.4987384901121112
X-intercept	0.01837315414798766	0.01850894096268858
Synthetic X-intercept	0.0	0.0
Elbow Point	0.5217867947322703	0.5283844364347974
Synthetic Elbow Point	0.5007451474615979	0.5015979212074365
JS Distance	0.04008094196088649	0.04302367299411442
Synthetic JS Distance	0.12973601083917569	0.13298968096921568
% Genome Enriched	40.96169014647004	40.21446329263057
Diff. Enrichment	7.014838305983267	7.095457505990815
CHANCE Divergence	0.05966320231350627	0.0603795616087285

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	0.5257289487308634	0.5134782133634901	0.374857874488584	0.3425428395733986	0.36689015230897654	0.3716271268121038	0.5037312885433273	0.5193517633425485	0.5206488894811536

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	0.38736824183936397	0.1958483073399561	0.19913382406849348	0.38985859309961585

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	0.026812840386302186	0.016996379972820663	0.019274499681203084	0.028132118388426983

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