QC Report

	general
Report generated at	2021-08-30 14:51:46
Title	hmg-20_RW12240_youngadult_1_12
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	13807128	15427464	16452922
Total Reads (QC-failed)	0	0	0
Duplicate Reads	0	0	0
Duplicate Reads (QC-failed)	0	0	0
Mapped Reads	13192832	14317333	16395759
Mapped Reads (QC-failed)	0	0	0
% Mapped Reads	95.6	92.80000000000001	99.7
Paired Reads	13807128	15427464	16452922
Paired Reads (QC-failed)	0	0	0
Read1	6903564	7713732	8226461
Read1 (QC-failed)	0	0	0
Read2	6903564	7713732	8226461
Read2 (QC-failed)	0	0	0
Properly Paired Reads	13105390	14172412	16300828
Properly Paired Reads (QC-failed)	0	0	0
% Properly Paired Reads	94.89999999999999	91.9	99.1
With itself	13182258	14303908	16385262
With itself (QC-failed)	0	0	0
Singletons	10574	13425	10497
Singletons (QC-failed)	0	0	0
% Singleton	0.1	0.1	0.1
Diff. Chroms	3642	4428	31291
Diff. Chroms (QC-failed)	0	0	0

Marking duplicates (filtered BAM)

	rep1	rep2	ctl1
Unpaired Reads	0	0	0
Paired Reads	5992635	6471749	7495598
Unmapped Reads	0	0	0
Unpaired Duplicate Reads	0	0	0
Paired Duplicate Reads	1148212	1485302	1278205
Paired Optical Duplicate Reads	66826	86644	82589
% Duplicate Reads	19.1604	22.950599999999998	17.0527

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	9688846	9972894	12434786
Total Reads (QC-failed)	0	0	0
Duplicate Reads	0	0	0
Duplicate Reads (QC-failed)	0	0	0
Mapped Reads	9688846	9972894	12434786
Mapped Reads (QC-failed)	0	0	0
% Mapped Reads	100.0	100.0	100.0
Paired Reads	9688846	9972894	12434786
Paired Reads (QC-failed)	0	0	0
Read1	4844423	4986447	6217393
Read1 (QC-failed)	0	0	0
Read2	4844423	4986447	6217393
Read2 (QC-failed)	0	0	0
Properly Paired Reads	9688846	9972894	12434786
Properly Paired Reads (QC-failed)	0	0	0
% Properly Paired Reads	100.0	100.0	100.0
With itself	9688846	9972894	12434786
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	5876124	6331600	7356785
Distinct Fragments	4759064	4896008	6109413
Positions with Two Read	783221	925100	927452
NRF = Distinct/Total	0.809898	0.773266	0.830446
PBC1 = OneRead/Distinct	0.803192	0.764128	0.823743
PBC2 = OneRead/TwoRead	4.880416	4.044081	5.426248

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	41713	999
N1	31505	519
N2	33007	475
Np	41608	1016
N optimal	41713	1016
N conservative	41713	999
Optimal Set	rep1_vs_rep2	pooled-pr1_vs_pooled-pr2
Conservative Set	rep1_vs_rep2	rep1_vs_rep2
Rescue Ratio	1.0025235531628534	1.017017017017017
Self Consistency Ratio	1.0476749722266308	1.0926315789473684
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	82228	87311

Top 300000 raw peaks from spp with FDR 0.01

Peak calling statistics

Peak region size

	rep1	rep2	idr_opt	overlap_opt
Min size	62.0	55.0	64.0	64.0
25 percentile	250.0	220.0	256.0	256.0
50 percentile (median)	250.0	220.0	256.0	256.0
75 percentile	250.0	220.0	256.0	256.0
Max size	280.0	318.0	339.0	339.0
Mean	249.92035559663375	219.9634295793199	246.5738188976378	255.77448277515404

Enrichment / Signal-to-noise ratio

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

	rep1	rep2
Number of Subsampled Reads	5554641	5992526
Estimated Fragment Length	125	130
Cross-correlation at Estimated Fragment Length	0.606431811784532	0.608386928556483
Phantom Peak	50	50
Cross-correlation at Phantom Peak	0.6064607	0.6084019
Argmin of Cross-correlation	1500	1500
Minimum of Cross-correlation	0.6019335	0.60359
NSC (Normalized Strand Cross-correlation coeff.)	1.007473	1.007947
RSC (Relative Strand Cross-correlation coeff.)	0.9936299	0.9968786

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	0.3947114329831993	0.3943493772868193
Synthetic AUC	0.49793523283783164	0.4979390631988593
X-intercept	0.019823735795284455	0.020025306047506717
Synthetic X-intercept	0.0	0.0
Elbow Point	0.5492549883648062	0.5474308773694483
Synthetic Elbow Point	0.4992187567770514	0.5028191347578155
JS Distance	0.05079694471614301	0.05051715500430097
Synthetic JS Distance	0.1683654721342655	0.17088191471917782
% Genome Enriched	42.04436142144947	41.69809965154291
Diff. Enrichment	10.879135855554322	10.713711994956537
CHANCE Divergence	0.09249700852606663	0.09108322130760682

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	0.38555592688747453	0.3752546652957507	0.40689894195883763	0.3997480771884115	0.39919334029942066	0.40373745148348145	0.49647548996172264	0.37816492779073924	0.3780379311369047

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	0.20553669207303119	0.17298468775331965	0.1666923362466301	0.2055516449714013

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	0.020327092108836757	0.0170797430364772	0.018759750178834748	0.020797345504517912

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