QC Report

	general
Report generated at	2020-03-28 20:51:39
Title	pho4_pbexo
Description	pb exo data for pho4
Pipeline version	v1.3.6
Pipeline type	tf
Genome	sacCer3
Aligner	bowtie2
Sequencing endedness	{'rep1': {'paired_end': True}, 'rep2': {'paired_end': True}, 'ctl1': {'paired_end': False}}
Peak caller	macs2

Alignment quality metrics

SAMstat (raw unfiltered BAM)

	rep1	rep2	ctl1
Total Reads	2056200	3011704	6055441
Total Reads (QC-failed)	0	0	0
Duplicate Reads	0	0	0
Duplicate Reads (QC-failed)	0	0	0
Mapped Reads	1696849	2021447	5615958
Mapped Reads (QC-failed)	0	0	0
% Mapped Reads	82.5	67.10000000000001	92.7
Paired Reads	2056200	3011704	0
Paired Reads (QC-failed)	0	0	0
Read1	1028100	1505852	0
Read1 (QC-failed)	0	0	0
Read2	1028100	1505852	0
Read2 (QC-failed)	0	0	0
Properly Paired Reads	1402884	1691938	0
Properly Paired Reads (QC-failed)	0	0	0
% Properly Paired Reads	68.2	56.2	0.0
With itself	1543754	1855304	0
With itself (QC-failed)	0	0	0
Singletons	153095	166143	0
Singletons (QC-failed)	0	0	0
% Singleton	7.3999999999999995	5.5	0.0
Diff. Chroms	74979	94153	0
Diff. Chroms (QC-failed)	0	0	0

Marking duplicates (filtered BAM)

	rep1	rep2	ctl1
Unpaired Reads	0	0	4971580
Paired Reads	588888	711623	0
Unmapped Reads	0	0	0
Unpaired Duplicate Reads	0	0	1717069
Paired Duplicate Reads	343825	430108	0
Paired Optical Duplicate Reads	0	0	0
% Duplicate Reads	58.3855	60.440400000000004	34.5377

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	490126	563030	3254511
Total Reads (QC-failed)	0	0	0
Duplicate Reads	0	0	0
Duplicate Reads (QC-failed)	0	0	0
Mapped Reads	490126	563030	3254511
Mapped Reads (QC-failed)	0	0	0
% Mapped Reads	100.0	100.0	100.0
Paired Reads	490126	563030	0
Paired Reads (QC-failed)	0	0	0
Read1	245063	281515	0
Read1 (QC-failed)	0	0	0
Read2	245063	281515	0
Read2 (QC-failed)	0	0	0
Properly Paired Reads	490126	563030	0
Properly Paired Reads (QC-failed)	0	0	0
% Properly Paired Reads	100.0	100.0	0.0
With itself	490126	563030	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	580410	701486	4962084
Distinct Fragments	241285	277223	3247815
Positions with Two Read	61711	70188	586396
NRF = Distinct/Total	0.415715	0.395194	0.654526
PBC1 = OneRead/Distinct	0.388677	0.355854	0.747173
PBC2 = OneRead/TwoRead	1.519697	1.405525	4.138296

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	2123	1202
N1	1558	767
N2	1668	906
Np	2095	1174
N optimal	2123	1202
N conservative	2123	1202
Optimal Set	rep1_vs_rep2	rep1_vs_rep2
Conservative Set	rep1_vs_rep2	rep1_vs_rep2
Rescue Ratio	1.013365155131265	1.0238500851788757
Self Consistency Ratio	1.0706033376123234	1.18122555410691
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	2615	2626

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	150.0	150.0	183.0	167.0
25 percentile	213.0	221.0	335.0	284.0
50 percentile (median)	285.0	292.0	418.0	342.0
75 percentile	382.0	390.0	527.0	459.5
Max size	2129.0	1864.0	2716.0	2716.0
Mean	318.45200764818355	328.9207920792079	467.1963394342762	397.0640602920396

Enrichment / Signal-to-noise ratio

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

	rep1	rep2
Number of Subsampled Reads	727725	863648
Estimated Fragment Length	90	130
Cross-correlation at Estimated Fragment Length	0.384077727569322	0.39698344981185
Phantom Peak	30	30
Cross-correlation at Phantom Peak	0.4443394	0.4702
Argmin of Cross-correlation	1500	1500
Minimum of Cross-correlation	0.3457825	0.3666965
NSC (Normalized Strand Cross-correlation coeff.)	1.11075	1.082594
RSC (Relative Strand Cross-correlation coeff.)	0.38856	0.2926177

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	0.27178150758430664	0.2725458177352572
Synthetic AUC	0.49502525315309126	0.49535773521780885
X-intercept	0.03283064996898792	0.03197555967265698
Synthetic X-intercept	0.0	0.0
Elbow Point	0.7045964114309113	0.7097791418185322
Synthetic Elbow Point	0.5005572772239844	0.5072580786268486
JS Distance	0.23547855672856374	0.23559118607904414
Synthetic JS Distance	0.3283494023144413	0.32851765987028325
% Genome Enriched	23.80663718679307	24.039077225091482
Diff. Enrichment	28.66395085125016	28.63485101893188
CHANCE Divergence	0.24899454988451486	0.2484645303572877

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	0.3682481647576338	0.37698346446903364	0.33747102797636536	0.3508184259509229	0.3387510099485028	0.35275332665515746	0.4004022196141882	0.374738881081697	0.3752563732490657

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	0.35625206522110686	0.3036280466655513	0.3183258440935652	0.3546777495451766

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	0.2790868589268826	0.21516712029151688	0.23906719002539828	0.27650794374242754

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