QC Report

	general
Report generated at	2022-12-20 14:21:00
Title	hmg-4_OP660_earlyembryonic_1_1
Description	ENCSR306UKE
Pipeline version	v1.3.6
Pipeline type	tf
Genome	WS245chr
Aligner	bwa
Sequencing endedness	{'rep1': {'paired_end': False}, 'rep2': {'paired_end': False}, 'ctl1': {'paired_end': False}}
Peak caller	spp

Alignment quality metrics

Marking duplicates (filtered BAM)

	rep1	rep2	ctl1
Unpaired Reads	13762396	20182600	19427277
Paired Reads	0	0	0
Unmapped Reads	0	0	0
Unpaired Duplicate Reads	1763751	2969548	2659107
Paired Duplicate Reads	0	0	0
Paired Optical Duplicate Reads	0	0	0
% Duplicate Reads	12.8157	14.713399999999998	13.6875

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	11998645	17213052	16768170
Total Reads (QC-failed)	0	0	0
Duplicate Reads	0	0	0
Duplicate Reads (QC-failed)	0	0	0
Mapped Reads	11998645	17213052	16768170
Mapped Reads (QC-failed)	0	0	0
% Mapped Reads	100.0	100.0	100.0
Paired Reads	0	0	0
Paired Reads (QC-failed)	0	0	0
Read1	0	0	0
Read1 (QC-failed)	0	0	0
Read2	0	0	0
Read2 (QC-failed)	0	0	0
Properly Paired Reads	0	0	0
Properly Paired Reads (QC-failed)	0	0	0
% Properly Paired Reads	0.0	0.0	0.0
With itself	0	0	0
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	13749743	20115701	19272170
Distinct Fragments	11992973	17195514	16745690
Positions with Two Read	1302450	2144878	1936756
NRF = Distinct/Total	0.872233	0.85483	0.868905
PBC1 = OneRead/Distinct	0.876048	0.855972	0.868694
PBC2 = OneRead/TwoRead	8.066659	6.862337	7.51095

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	23383	1362
N1	43231	1767
N2	10676	709
Np	52147	3209
N optimal	52147	3209
N conservative	23383	1362
Optimal Set	pooled-pr1_vs_pooled-pr2	pooled-pr1_vs_pooled-pr2
Conservative Set	rep1_vs_rep2	rep1_vs_rep2
Rescue Ratio	2.2301244493863064	2.3560939794419973
Self Consistency Ratio	4.0493630573248405	2.4922425952045133
Reproducibility Test	fail	fail

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	59807	27558

Top 300000 raw peaks from spp with FDR 0.01

Peak calling statistics

Peak region size

	rep1	rep2	idr_opt	overlap_opt
Min size	128.0	158.0	136.0	136.0
25 percentile	510.0	630.0	544.0	544.0
50 percentile (median)	510.0	630.0	544.0	544.0
75 percentile	510.0	630.0	544.0	544.0
Max size	759.0	791.0	885.0	885.0
Mean	508.88394335111275	628.776761738878	519.9211592396385	542.4042610313153

Enrichment / Signal-to-noise ratio

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

	rep1	rep2
Number of Subsampled Reads	13749743	15000000
Estimated Fragment Length	235	220
Cross-correlation at Estimated Fragment Length	0.82294552231168	0.836283974747981
Phantom Peak	50	50
Cross-correlation at Phantom Peak	0.8125595	0.8359166
Argmin of Cross-correlation	1500	1500
Minimum of Cross-correlation	0.8040153	0.8311738
NSC (Normalized Strand Cross-correlation coeff.)	1.023545	1.006148
RSC (Relative Strand Cross-correlation coeff.)	2.215552	1.077456

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	0.348206661455733	0.4080879811962727
Synthetic AUC	0.4974133938514791	0.4978400370986824
X-intercept	0.028981023308176125	0.028505957287971846
Synthetic X-intercept	0.0	0.0
Elbow Point	0.5479627053213383	0.46888018156304706
Synthetic Elbow Point	0.5004396683685087	0.5020913486718602
JS Distance	0.13163115377281412	0.04362861652473152
Synthetic JS Distance	0.21410687160880643	0.13942983469982248
% Genome Enriched	39.06879075737101	35.350900130343746
Diff. Enrichment	17.036750465390288	6.543900055977192
CHANCE Divergence	0.14513777381779486	0.056112584662324744

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	0.41080497006120276	0.20967368250557775	0.4300106862057602	0.1845218384281881	0.4310935469041335	0.17783644643611138	0.40890750030715434	0.42328782120094494	0.4258440865603969

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	0.16087521378850397	0.32364429483495843	0.08998084709207874	0.3265234813300987

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	0.018521245102603934	0.02774504954517781	0.010982538134434266	0.03911936372611286

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