QC Report

	general
Report generated at	2021-03-26 10:15:57
Title	myc_hela_1
Description	chipseq of myc_hela_1
Pipeline version	v1.3.6
Pipeline type	tf
Genome	hg38
Aligner	bowtie2
Sequencing endedness	{'rep1': {'paired_end': False}, 'rep2': {'paired_end': False}, 'ctl1': {'paired_end': False}}
Peak caller	macs2

Alignment quality metrics

SAMstat (raw unfiltered BAM)

	rep1	rep2	ctl1
Total Reads	10570866	8433611	19922945
Total Reads (QC-failed)	0	0	0
Duplicate Reads	0	0	0
Duplicate Reads (QC-failed)	0	0	0
Mapped Reads	9027123	6835126	17960881
Mapped Reads (QC-failed)	0	0	0
% Mapped Reads	85.39999999999999	81.0	90.2
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

Marking duplicates (filtered BAM)

	rep1	rep2	ctl1
Unpaired Reads	6866184	5051942	12786426
Paired Reads	0	0	0
Unmapped Reads	0	0	0
Unpaired Duplicate Reads	234087	459274	1489721
Paired Duplicate Reads	0	0	0
Paired Optical Duplicate Reads	0	0	0
% Duplicate Reads	3.4093	9.091000000000001	11.6508

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	6866184	5051942	12786426
Total Reads (QC-failed)	0	0	0
Duplicate Reads	0	0	0
Duplicate Reads (QC-failed)	0	0	0
Mapped Reads	6866184	5051942	12786426
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	6842684	5040017	12714374
Distinct Fragments	6619706	4584997	11305381
Positions with Two Read	203189	376868	1377145
NRF = Distinct/Total	0.967414	0.909719	0.889181
PBC1 = OneRead/Distinct	0.967913	0.90984	0.87708
PBC2 = OneRead/TwoRead	31.533695	11.069165	7.200201

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	50040	12176
N1	70758	30899
N2	95951	24242
Np	47971	15157
N optimal	50040	15157
N conservative	50040	12176
Optimal Set	rep1_vs_rep2	pooled-pr1_vs_pooled-pr2
Conservative Set	rep1_vs_rep2	rep1_vs_rep2
Rescue Ratio	1.0431302245106417	1.2448258869908015
Self Consistency Ratio	1.356044546199723	1.274606055605973
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	183652	499277

Top 500000 raw peaks from macs2 with p-val threshold 0.01

Peak calling statistics

Peak region size

	rep1	rep2	idr_opt	overlap_opt
Min size	110.0	75.0	92.0	92.0
25 percentile	110.0	75.0	230.0	153.0
50 percentile (median)	110.0	75.0	310.0	212.0
75 percentile	174.0	106.0	421.0	304.0
Max size	1576.0	962.0	1874.0	1874.0
Mean	162.28428767451484	97.68276527859284	345.3841129511117	248.84680255795362

Enrichment / Signal-to-noise ratio

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

	rep1	rep2
Number of Subsampled Reads	6842684	5040017
Estimated Fragment Length	110	75
Cross-correlation at Estimated Fragment Length	0.135457915589408	0.0859181381066594
Phantom Peak	35	35
Cross-correlation at Phantom Peak	0.122905	0.0800039
Argmin of Cross-correlation	1500	1500
Minimum of Cross-correlation	0.08405237	0.05568788
NSC (Normalized Strand Cross-correlation coeff.)	1.611589	1.542852
RSC (Relative Strand Cross-correlation coeff.)	1.32309	1.243224

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	0.1339671803524985	0.12884703343576343
Synthetic AUC	0.4742624318332945	0.4699541992997388
X-intercept	0.5034168656006206	0.5697488433839966
Synthetic X-intercept	2.486435632823678e-12	5.486593236858868e-09
Elbow Point	0.7675731657089846	0.5923373440017914
Synthetic Elbow Point	0.5367836849274152	0.5425209307202643
JS Distance	0.15938491955479778	0.1387296280677433
Synthetic JS Distance	0.3267478299887218	0.27016054910590304
% Genome Enriched	22.4541452237857	40.56553224329522
Diff. Enrichment	36.394004801814575	41.16011890648135
CHANCE Divergence	0.31786963288510955	0.46355570479132496

Peak enrichment

Fraction of reads in peaks (FRiP)

FRiP for macs2 raw peaks

	rep1	rep2	rep1-pr1	rep2-pr1	rep1-pr2	rep2-pr2	pooled	pooled-pr1	pooled-pr2
Fraction of Reads in Peaks	0.23666755799145495	0.3327053240120334	0.35025452274509394	0.37446312724888764	0.35055687409483927	0.3749995546267158	0.24190271188608006	0.20014925165248296	0.20062097010889127

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	0.14361267870468897	0.1674650723021696	0.13091757585498803	0.14593099619856342

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	0.0774740928229824	0.13663120592165895	0.08724526132722822	0.08833846864851068

For macs2 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