QC Report

	general
Report generated at	2021-10-25 10:28:00
Title	sma-3_CS152_L4larva_1_1
Description	mkudron
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	15442822	14400636	12232442
Total Reads (QC-failed)	0	0	0
Duplicate Reads	0	0	0
Duplicate Reads (QC-failed)	0	0	0
Mapped Reads	15330775	14310293	12152909
Mapped Reads (QC-failed)	0	0	0
% Mapped Reads	99.3	99.4	99.3
Paired Reads	15442822	14400636	12232442
Paired Reads (QC-failed)	0	0	0
Read1	7721411	7200318	6116221
Read1 (QC-failed)	0	0	0
Read2	7721411	7200318	6116221
Read2 (QC-failed)	0	0	0
Properly Paired Reads	15260032	14259962	12092074
Properly Paired Reads (QC-failed)	0	0	0
% Properly Paired Reads	98.8	99.0	98.9
With itself	15311428	14296776	12140392
With itself (QC-failed)	0	0	0
Singletons	19347	13517	12517
Singletons (QC-failed)	0	0	0
% Singleton	0.1	0.1	0.1
Diff. Chroms	18204	8044	15098
Diff. Chroms (QC-failed)	0	0	0

Marking duplicates (filtered BAM)

	rep1	rep2	ctl1
Unpaired Reads	0	0	0
Paired Reads	6977117	6516007	5529240
Unmapped Reads	0	0	0
Unpaired Duplicate Reads	0	0	0
Paired Duplicate Reads	792437	793513	592568
Paired Optical Duplicate Reads	82551	78260	68141
% Duplicate Reads	11.3577	12.1779	10.717

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	12369360	11444988	9873344
Total Reads (QC-failed)	0	0	0
Duplicate Reads	0	0	0
Duplicate Reads (QC-failed)	0	0	0
Mapped Reads	12369360	11444988	9873344
Mapped Reads (QC-failed)	0	0	0
% Mapped Reads	100.0	100.0	100.0
Paired Reads	12369360	11444988	9873344
Paired Reads (QC-failed)	0	0	0
Read1	6184680	5722494	4936672
Read1 (QC-failed)	0	0	0
Read2	6184680	5722494	4936672
Read2 (QC-failed)	0	0	0
Properly Paired Reads	12369360	11444988	9873344
Properly Paired Reads (QC-failed)	0	0	0
% Properly Paired Reads	100.0	100.0	100.0
With itself	12369360	11444988	9873344
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	6944437	6473481	5477798
Distinct Fragments	6161618	5689538	4894658
Positions with Two Read	638108	626114	484236
NRF = Distinct/Total	0.887274	0.878899	0.893545
PBC1 = OneRead/Distinct	0.885323	0.876882	0.89146
PBC2 = OneRead/TwoRead	8.548747	7.968281	9.010875

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	85979	796
N1	37386	198
N2	30482	352
Np	85672	824
N optimal	85979	824
N conservative	85979	796
Optimal Set	rep1_vs_rep2	pooled-pr1_vs_pooled-pr2
Conservative Set	rep1_vs_rep2	rep1_vs_rep2
Rescue Ratio	1.0035834344943506	1.035175879396985
Self Consistency Ratio	1.2264943245193884	1.7777777777777777
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	128649	129781

Top 300000 raw peaks from spp with FDR 0.01

Peak calling statistics

Peak region size

	rep1	rep2	idr_opt	overlap_opt
Min size	122.0	79.0	88.0	88.0
25 percentile	336.0	316.0	350.0	350.0
50 percentile (median)	336.0	316.0	350.0	350.0
75 percentile	336.0	316.0	350.0	350.0
Max size	336.0	316.0	350.0	350.0
Mean	335.994698753974	315.98237800602556	345.2621359223301	349.9496853882925

Enrichment / Signal-to-noise ratio

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

	rep1	rep2
Number of Subsampled Reads	6542106	6091820
Estimated Fragment Length	195	180
Cross-correlation at Estimated Fragment Length	0.673622768735163	0.654708661328661
Phantom Peak	50	50
Cross-correlation at Phantom Peak	0.6737317	0.655059
Argmin of Cross-correlation	1500	1500
Minimum of Cross-correlation	0.6696194	0.6498769
NSC (Normalized Strand Cross-correlation coeff.)	1.005979	1.007435
RSC (Relative Strand Cross-correlation coeff.)	0.9735207	0.9323983

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	0.41741333328217334	0.41344165848427245
Synthetic AUC	0.4981879937478838	0.4981148819902422
X-intercept	0.01859368078253894	0.018755312907821797
Synthetic X-intercept	0.0	0.0
Elbow Point	0.5110308936859708	0.5187612992828323
Synthetic Elbow Point	0.5023087596130391	0.4998887865205286
JS Distance	0.028304294281024996	0.026111518501140372
Synthetic JS Distance	0.12862394229768195	0.1359136497976675
% Genome Enriched	44.74376319496825	44.01482226452594
Diff. Enrichment	8.035941814140529	8.460616121428476
CHANCE Divergence	0.06832850889833157	0.07192442155806611

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	0.5758876772929238	0.5687824224892154	0.4220250683948078	0.3881127704109432	0.4201706474708473	0.37471668821321613	0.5722888571209256	0.5725530675876577	0.5729304031334387

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	0.4137961702751635	0.20842679006836248	0.17060000412407597	0.41263888475972554

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	0.007804748633050966	0.002323968257048061	0.0043921409091909926	0.00810104059955788

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