QC Report

	general
Report generated at	2024-01-11 14:20:43
Title	sptf-3_RW12349_youngadult_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	23523638	28704574	29437072
Total Reads (QC-failed)	0	0	0
Duplicate Reads	0	0	0
Duplicate Reads (QC-failed)	0	0	0
Mapped Reads	20826496	28250493	29417076
Mapped Reads (QC-failed)	0	0	0
% Mapped Reads	88.5	98.4	99.9
Paired Reads	23523638	28704574	29437072
Paired Reads (QC-failed)	0	0	0
Read1	11761819	14352287	14718536
Read1 (QC-failed)	0	0	0
Read2	11761819	14352287	14718536
Read2 (QC-failed)	0	0	0
Properly Paired Reads	20733954	28142204	29211958
Properly Paired Reads (QC-failed)	0	0	0
% Properly Paired Reads	88.1	98.0	99.2
With itself	20813448	28243318	29409368
With itself (QC-failed)	0	0	0
Singletons	13048	7175	7708
Singletons (QC-failed)	0	0	0
% Singleton	0.1	0.0	0.0
Diff. Chroms	14386	19159	86779
Diff. Chroms (QC-failed)	0	0	0

Marking duplicates (filtered BAM)

	rep1	rep2	ctl1
Unpaired Reads	0	0	0
Paired Reads	9355187	12894873	12973507
Unmapped Reads	0	0	0
Unpaired Duplicate Reads	0	0	0
Paired Duplicate Reads	1366797	2632390	1383630
Paired Optical Duplicate Reads	719447	760451	709925
% Duplicate Reads	14.610000000000001	20.414199999999997	10.665

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	15976780	20524966	23179754
Total Reads (QC-failed)	0	0	0
Duplicate Reads	0	0	0
Duplicate Reads (QC-failed)	0	0	0
Mapped Reads	15976780	20524966	23179754
Mapped Reads (QC-failed)	0	0	0
% Mapped Reads	100.0	100.0	100.0
Paired Reads	15976780	20524966	23179754
Paired Reads (QC-failed)	0	0	0
Read1	7988390	10262483	11589877
Read1 (QC-failed)	0	0	0
Read2	7988390	10262483	11589877
Read2 (QC-failed)	0	0	0
Properly Paired Reads	15976780	20524966	23179754
Properly Paired Reads (QC-failed)	0	0	0
% Properly Paired Reads	100.0	100.0	100.0
With itself	15976780	20524966	23179754
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	9327186	12847074	12695085
Distinct Fragments	7956017	10314940	11385699
Positions with Two Read	920264	1621445	998564
NRF = Distinct/Total	0.852992	0.802902	0.896859
PBC1 = OneRead/Distinct	0.861263	0.804397	0.900392
PBC2 = OneRead/TwoRead	7.445929	5.117228	10.266335

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	36268	4513
N1	36799	3596
N2	34075	4294
Np	38361	5236
N optimal	38361	5236
N conservative	36268	4513
Optimal Set	pooled-pr1_vs_pooled-pr2	pooled-pr1_vs_pooled-pr2
Conservative Set	rep1_vs_rep2	rep1_vs_rep2
Rescue Ratio	1.057709275394287	1.1602038555284733
Self Consistency Ratio	1.0799413059427734	1.1941045606229144
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	65037	53867

Top 300000 raw peaks from spp with FDR 0.01

Peak calling statistics

Peak region size

	rep1	rep2	idr_opt	overlap_opt
Min size	94.0	94.0	104.0	104.0
25 percentile	376.0	376.0	264.0	416.0
50 percentile (median)	376.0	376.0	416.0	416.0
75 percentile	376.0	376.0	416.0	416.0
Max size	2046.0	1739.0	3417.0	3417.0
Mean	373.5545151221612	371.671412924425	397.4749809014515	411.6892416777456

Enrichment / Signal-to-noise ratio

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

	rep1	rep2
Number of Subsampled Reads	8398519	11826848
Estimated Fragment Length	185	180
Cross-correlation at Estimated Fragment Length	0.703582815770636	0.753832274175968
Phantom Peak	50	55
Cross-correlation at Phantom Peak	0.694064	0.7406686
Argmin of Cross-correlation	1500	1500
Minimum of Cross-correlation	0.6834015	0.7261302
NSC (Normalized Strand Cross-correlation coeff.)	1.029531	1.03815
RSC (Relative Strand Cross-correlation coeff.)	1.892736	1.905439

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	0.35198023028779823	0.3342195221028437
Synthetic AUC	0.4986591641379292	0.49882368871579835
X-intercept	0.01768206234150043	0.017636134906847182
Synthetic X-intercept	0.0	0.0
Elbow Point	0.667812855688013	0.6878711635615727
Synthetic Elbow Point	0.5017422642214202	0.49934848646302626
JS Distance	0.11858874249539983	0.13876960335682925
Synthetic JS Distance	0.22598380555006897	0.25316918972592056
% Genome Enriched	18.783322350685918	19.972842908205035
Diff. Enrichment	12.934246514529836	15.544242031032486
CHANCE Divergence	0.11883129794929707	0.13945612351033754

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	0.38542422190203535	0.3709447801277722	0.4139234564161239	0.3940145485245093	0.41516288012978836	0.3923789586183927	0.35446060580225397	0.37189769651579424	0.3708918127309205

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	0.2892875590115607	0.27827522191580534	0.29399887921860623	0.3014068970837724

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	0.09058284499596266	0.06981500652822409	0.09257584153854384	0.09930363331112983

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