QC Report

	general
Report generated at	2021-03-25 20:21:17
Title	mxi_gm12878
Description	chipseq of mxi_gm12878
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}, 'ctl2': {'paired_end': False}}
Peak caller	macs2

Alignment quality metrics

SAMstat (raw unfiltered BAM)

	rep1	rep2	ctl1	ctl2
Total Reads	39422373	23205307	5526289	6211790
Total Reads (QC-failed)	0	0	0	0
Duplicate Reads	0	0	0	0
Duplicate Reads (QC-failed)	0	0	0	0
Mapped Reads	36185709	22757273	4962259	5760784
Mapped Reads (QC-failed)	0	0	0	0
% Mapped Reads	91.8	98.1	89.8	92.7
Paired Reads	0	0	0	0
Paired Reads (QC-failed)	0	0	0	0
Read1	0	0	0	0
Read1 (QC-failed)	0	0	0	0
Read2	0	0	0	0
Read2 (QC-failed)	0	0	0	0
Properly Paired Reads	0	0	0	0
Properly Paired Reads (QC-failed)	0	0	0	0
% Properly Paired Reads	0.0	0.0	0.0	0.0
With itself	0	0	0	0
With itself (QC-failed)	0	0	0	0
Singletons	0	0	0	0
Singletons (QC-failed)	0	0	0	0
% Singleton	0.0	0.0	0.0	0.0
Diff. Chroms	0	0	0	0
Diff. Chroms (QC-failed)	0	0	0	0

Marking duplicates (filtered BAM)

	rep1	rep2	ctl1	ctl2
Unpaired Reads	30199564	18683337	3930750	4510303
Paired Reads	0	0	0	0
Unmapped Reads	0	0	0	0
Unpaired Duplicate Reads	4140307	506528	126940	127662
Paired Duplicate Reads	0	0	0	0
Paired Optical Duplicate Reads	0	0	0	0
% Duplicate Reads	13.7098	2.7111	3.2294000000000005	2.8305000000000002

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	ctl2
Total Reads	30199564	18683337	3930750	4510303
Total Reads (QC-failed)	0	0	0	0
Duplicate Reads	0	0	0	0
Duplicate Reads (QC-failed)	0	0	0	0
Mapped Reads	30199564	18683337	3930750	4510303
Mapped Reads (QC-failed)	0	0	0	0
% Mapped Reads	100.0	100.0	100.0	100.0
Paired Reads	0	0	0	0
Paired Reads (QC-failed)	0	0	0	0
Read1	0	0	0	0
Read1 (QC-failed)	0	0	0	0
Read2	0	0	0	0
Read2 (QC-failed)	0	0	0	0
Properly Paired Reads	0	0	0	0
Properly Paired Reads (QC-failed)	0	0	0	0
% Properly Paired Reads	0.0	0.0	0.0	0.0
With itself	0	0	0	0
With itself (QC-failed)	0	0	0	0
Singletons	0	0	0	0
Singletons (QC-failed)	0	0	0	0
% Singleton	0.0	0.0	0.0	0.0
Diff. Chroms	0	0	0	0
Diff. Chroms (QC-failed)	0	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	ctl2
Total Fragments	30107202	18647033	3898040	4488260
Distinct Fragments	26038463	18160120	3789354	4370528
Positions with Two Read	3191045	443510	101113	110480
NRF = Distinct/Total	0.864858	0.973888	0.972118	0.973769
PBC1 = OneRead/Distinct	0.862279	0.974661	0.972505	0.974007
PBC2 = OneRead/TwoRead	7.036068	39.908807	36.446016	38.531164

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	14997	4484
N1	16813	5368
N2	15218	3896
Np	16424	6201
N optimal	16424	6201
N conservative	14997	4484
Optimal Set	pooled-pr1_vs_pooled-pr2	pooled-pr1_vs_pooled-pr2
Conservative Set	rep1_vs_rep2	rep1_vs_rep2
Rescue Ratio	1.0951523638060945	1.3829170383586085
Self Consistency Ratio	1.1048100933105534	1.37782340862423
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	22863	24131

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	115.0	120.0	118.0	118.0
25 percentile	145.0	145.0	257.0	179.0
50 percentile (median)	202.0	205.0	349.0	246.0
75 percentile	302.0	320.0	489.0	361.0
Max size	2660.0	1676.0	1631.0	1631.0
Mean	251.59012378078117	262.56006796237205	397.45121754555714	297.13041889917196

Enrichment / Signal-to-noise ratio

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

	rep1	rep2
Number of Subsampled Reads	15000000	15000000
Estimated Fragment Length	115	120
Cross-correlation at Estimated Fragment Length	0.171634382513905	0.180697619740161
Phantom Peak	40	35
Cross-correlation at Phantom Peak	0.1666848	0.1775928
Argmin of Cross-correlation	1500	1500
Minimum of Cross-correlation	0.1520085	0.1640724
NSC (Normalized Strand Cross-correlation coeff.)	1.12911	1.101328
RSC (Relative Strand Cross-correlation coeff.)	1.337252	1.229635

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	0.25852039587110154	0.25676869693686016
Synthetic AUC	0.48936194466133576	0.4864762010230293
X-intercept	0.1368140304857568	0.16658602709489612
Synthetic X-intercept	6.752491954014152e-75	5.468622747603961e-46
Elbow Point	0.5812347051390733	0.6419423073847187
Synthetic Elbow Point	0.4911815098483564	0.49033168194240007
JS Distance	0.1566341310364831	0.1504103074384355
Synthetic JS Distance	0.29238798135851046	0.2701174349843123
% Genome Enriched	33.59651157211177	33.88361510532461
Diff. Enrichment	18.457393777419774	21.331338633732695
CHANCE Divergence	0.15916348058447305	0.18374075769064027

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	0.04672696599195936	0.04622107924296393	0.052835663455273726	0.05257122683323505	0.052903147873260684	0.05267999248100018	0.042942725514592515	0.04551002311605804	0.04558653435045793

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	0.038631565667512247	0.04193428090551241	0.03963585305986827	0.040165905865529546

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	0.02167117700318154	0.024519294384514955	0.019824777554459355	0.025703936842864542

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