QC Report

	general
Report generated at	2021-08-28 13:11:14
Title	ceh-40_OP232_youngadult_1_5
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	13688108	13341646	14253916
Total Reads (QC-failed)	0	0	0
Duplicate Reads	0	0	0
Duplicate Reads (QC-failed)	0	0	0
Mapped Reads	13433980	13240026	14195470
Mapped Reads (QC-failed)	0	0	0
% Mapped Reads	98.1	99.2	99.6
Paired Reads	13688108	13341646	14253916
Paired Reads (QC-failed)	0	0	0
Read1	6844054	6670823	7126958
Read1 (QC-failed)	0	0	0
Read2	6844054	6670823	7126958
Read2 (QC-failed)	0	0	0
Properly Paired Reads	13381854	13195130	14130884
Properly Paired Reads (QC-failed)	0	0	0
% Properly Paired Reads	97.8	98.9	99.1
With itself	13420726	13226198	14180654
With itself (QC-failed)	0	0	0
Singletons	13254	13828	14816
Singletons (QC-failed)	0	0	0
% Singleton	0.1	0.1	0.1
Diff. Chroms	3037	2789	15805
Diff. Chroms (QC-failed)	0	0	0

Marking duplicates (filtered BAM)

	rep1	rep2	ctl1
Unpaired Reads	0	0	0
Paired Reads	6100444	6014204	6478621
Unmapped Reads	0	0	0
Unpaired Duplicate Reads	0	0	0
Paired Duplicate Reads	1153188	1037726	959360
Paired Optical Duplicate Reads	116255	108791	113704
% Duplicate Reads	18.9033	17.2546	14.8081

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	9894512	9952956	11038522
Total Reads (QC-failed)	0	0	0
Duplicate Reads	0	0	0
Duplicate Reads (QC-failed)	0	0	0
Mapped Reads	9894512	9952956	11038522
Mapped Reads (QC-failed)	0	0	0
% Mapped Reads	100.0	100.0	100.0
Paired Reads	9894512	9952956	11038522
Paired Reads (QC-failed)	0	0	0
Read1	4947256	4976478	5519261
Read1 (QC-failed)	0	0	0
Read2	4947256	4976478	5519261
Read2 (QC-failed)	0	0	0
Properly Paired Reads	9894512	9952956	11038522
Properly Paired Reads (QC-failed)	0	0	0
% Properly Paired Reads	100.0	100.0	100.0
With itself	9894512	9952956	11038522
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	6028573	5943441	6416996
Distinct Fragments	4905504	4932142	5475618
Positions with Two Read	806632	754216	730501
NRF = Distinct/Total	0.813709	0.829846	0.853299
PBC1 = OneRead/Distinct	0.805737	0.822802	0.84842
PBC2 = OneRead/TwoRead	4.900063	5.380654	6.359507

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	57135	713
N1	34339	296
N2	32063	287
Np	76859	867
N optimal	76859	867
N conservative	57135	713
Optimal Set	pooled-pr1_vs_pooled-pr2	pooled-pr1_vs_pooled-pr2
Conservative Set	rep1_vs_rep2	rep1_vs_rep2
Rescue Ratio	1.3452174674017678	1.2159887798036466
Self Consistency Ratio	1.0709852477934068	1.0313588850174216
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	138973	88815

Top 300000 raw peaks from spp with FDR 0.01

Peak calling statistics

Peak region size

	rep1	rep2	idr_opt	overlap_opt
Min size	52.0	60.0	60.0	60.0
25 percentile	210.0	230.0	240.0	240.0
50 percentile (median)	210.0	230.0	240.0	240.0
75 percentile	210.0	230.0	240.0	240.0
Max size	235.0	236.0	277.0	277.0
Mean	209.98991890511107	229.97476777571356	235.86851211072664	239.95414980678905

Enrichment / Signal-to-noise ratio

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

	rep1	rep2
Number of Subsampled Reads	5664433	5579472
Estimated Fragment Length	125	130
Cross-correlation at Estimated Fragment Length	0.610771657436212	0.61420863000586
Phantom Peak	50	50
Cross-correlation at Phantom Peak	0.6109159	0.614393
Argmin of Cross-correlation	1500	1500
Minimum of Cross-correlation	0.6066594	0.6102426
NSC (Normalized Strand Cross-correlation coeff.)	1.006779	1.006499
RSC (Relative Strand Cross-correlation coeff.)	0.9661209	0.9555717

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	0.40076702119206065	0.4011126123933545
Synthetic AUC	0.4979453822072131	0.49795812684262986
X-intercept	0.01974955291562879	0.019547965766111005
Synthetic X-intercept	0.0	0.0
Elbow Point	0.5236515616018395	0.5262163089991697
Synthetic Elbow Point	0.49878256977956315	0.5018652154421872
JS Distance	0.04578728083429794	0.045432992037914864
Synthetic JS Distance	0.15277385834389906	0.15239301906295746
% Genome Enriched	43.65818803091269	42.82609216324966
Diff. Enrichment	10.448221537656256	10.408302184499963
CHANCE Divergence	0.08887211961224034	0.0884923243428227

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	0.5168682396868082	0.38348898558378036	0.40627046589058663	0.3946029702130704	0.4077343076646933	0.3982171728680404	0.5217697290153082	0.5170975965296933	0.5172837159883568

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	0.24800579096537653	0.15871505335482944	0.15767978879842331	0.31596503896617945

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	0.013720792999892983	0.010287217803161995	0.010036415312194688	0.015327597454748385

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