QC Report

	general
Report generated at	2021-09-13 23:49:38
Title	ztf-29_RW12283_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	11661080	19627074	21273754
Total Reads (QC-failed)	0	0	0
Duplicate Reads	0	0	0
Duplicate Reads (QC-failed)	0	0	0
Mapped Reads	11642939	19561848	21230083
Mapped Reads (QC-failed)	0	0	0
% Mapped Reads	99.8	99.7	99.8
Paired Reads	11661080	19627074	21273754
Paired Reads (QC-failed)	0	0	0
Read1	5830540	9813537	10636877
Read1 (QC-failed)	0	0	0
Read2	5830540	9813537	10636877
Read2 (QC-failed)	0	0	0
Properly Paired Reads	11529954	19397282	20893322
Properly Paired Reads (QC-failed)	0	0	0
% Properly Paired Reads	98.9	98.8	98.2
With itself	11634322	19538530	21198098
With itself (QC-failed)	0	0	0
Singletons	8617	23318	31985
Singletons (QC-failed)	0	0	0
% Singleton	0.1	0.1	0.2
Diff. Chroms	5609	7758	16771
Diff. Chroms (QC-failed)	0	0	0

Marking duplicates (filtered BAM)

	rep1	rep2	ctl1
Unpaired Reads	0	0	0
Paired Reads	5169967	8764797	9548468
Unmapped Reads	0	0	0
Unpaired Duplicate Reads	0	0	0
Paired Duplicate Reads	830513	979094	1439592
Paired Optical Duplicate Reads	40481	95418	73595
% Duplicate Reads	16.0642	11.1708	15.0767

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	8678908	15571406	16217752
Total Reads (QC-failed)	0	0	0
Duplicate Reads	0	0	0
Duplicate Reads (QC-failed)	0	0	0
Mapped Reads	8678908	15571406	16217752
Mapped Reads (QC-failed)	0	0	0
% Mapped Reads	100.0	100.0	100.0
Paired Reads	8678908	15571406	16217752
Paired Reads (QC-failed)	0	0	0
Read1	4339454	7785703	8108876
Read1 (QC-failed)	0	0	0
Read2	4339454	7785703	8108876
Read2 (QC-failed)	0	0	0
Properly Paired Reads	8678908	15571406	16217752
Properly Paired Reads (QC-failed)	0	0	0
% Properly Paired Reads	100.0	100.0	100.0
With itself	8678908	15571406	16217752
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	5151853	8731968	9504582
Distinct Fragments	4325358	7763554	8074815
Positions with Two Read	623179	802154	1098423
NRF = Distinct/Total	0.839573	0.889096	0.849571
PBC1 = OneRead/Distinct	0.834054	0.886541	0.844761
PBC2 = OneRead/TwoRead	5.788995	8.580284	6.210078

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	36397	814
N1	28276	466
N2	32183	802
Np	36628	1114
N optimal	36628	1114
N conservative	36397	814
Optimal Set	pooled-pr1_vs_pooled-pr2	pooled-pr1_vs_pooled-pr2
Conservative Set	rep1_vs_rep2	rep1_vs_rep2
Rescue Ratio	1.0063466769239222	1.3685503685503686
Self Consistency Ratio	1.1381737162257746	1.7210300429184548
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	69556	86172

Top 300000 raw peaks from spp with FDR 0.01

Peak calling statistics

Peak region size

	rep1	rep2	idr_opt	overlap_opt
Min size	66.0	62.0	69.0	69.0
25 percentile	264.0	250.0	276.0	276.0
50 percentile (median)	264.0	250.0	276.0	276.0
75 percentile	264.0	250.0	276.0	276.0
Max size	264.0	250.0	279.0	279.0
Mean	263.9249525562137	249.89605672376177	260.8662477558348	275.53431800808124

Enrichment / Signal-to-noise ratio

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

	rep1	rep2
Number of Subsampled Reads	4883118	8245820
Estimated Fragment Length	130	145
Cross-correlation at Estimated Fragment Length	0.586896759955007	0.725914428189643
Phantom Peak	50	50
Cross-correlation at Phantom Peak	0.5871425	0.7257589
Argmin of Cross-correlation	1500	1500
Minimum of Cross-correlation	0.5822846	0.7204275
NSC (Normalized Strand Cross-correlation coeff.)	1.007921	1.007616
RSC (Relative Strand Cross-correlation coeff.)	0.9494196	1.029173

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	0.4039759566360114	0.41179089622179366
Synthetic AUC	0.4978289937689555	0.4983729384221357
X-intercept	0.01978144389168816	0.019302329295461985
Synthetic X-intercept	0.0	0.0
Elbow Point	0.5218276624797374	0.5171363320583562
Synthetic Elbow Point	0.5033535540461573	0.49963552225029795
JS Distance	0.04991583461503015	0.03910880069282916
Synthetic JS Distance	0.13899866988996828	0.13139999043928396
% Genome Enriched	42.73083341984014	42.95222428951298
Diff. Enrichment	9.361096021931125	7.882573143581062
CHANCE Divergence	0.07957971346897241	0.06699134646995933

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	0.34524458607004477	0.3758774255837912	0.4102430858813113	0.3628052132472542	0.4109848842734593	0.3652354020228362	0.5263802769729085	0.3520222994207581	0.3456701917896974

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	0.18078273130813893	0.1557550788647604	0.15850771600201036	0.18312187627756077

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	0.00898516200656206	0.006470053605822299	0.008976003836776204	0.011443934292974516

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