QC Report

	general
Report generated at	2023-07-21 16:58:32
Title	sta-1_RW12332_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	14703012	7451824	11894522
Total Reads (QC-failed)	0	0	0
Duplicate Reads	0	0	0
Duplicate Reads (QC-failed)	0	0	0
Mapped Reads	14621708	7412917	11835242
Mapped Reads (QC-failed)	0	0	0
% Mapped Reads	99.4	99.5	99.5
Paired Reads	14703012	7451824	11894522
Paired Reads (QC-failed)	0	0	0
Read1	7351506	3725912	5947261
Read1 (QC-failed)	0	0	0
Read2	7351506	3725912	5947261
Read2 (QC-failed)	0	0	0
Properly Paired Reads	14538582	7372502	11729092
Properly Paired Reads (QC-failed)	0	0	0
% Properly Paired Reads	98.9	98.9	98.6
With itself	14601646	7402512	11818562
With itself (QC-failed)	0	0	0
Singletons	20062	10405	16680
Singletons (QC-failed)	0	0	0
% Singleton	0.1	0.1	0.1
Diff. Chroms	8475	4968	22231
Diff. Chroms (QC-failed)	0	0	0

Marking duplicates (filtered BAM)

	rep1	rep2	ctl1
Unpaired Reads	0	0	0
Paired Reads	6695121	3393341	5403722
Unmapped Reads	0	0	0
Unpaired Duplicate Reads	0	0	0
Paired Duplicate Reads	809186	331075	602050
Paired Optical Duplicate Reads	55765	22062	46468
% Duplicate Reads	12.0862	9.7566	11.141399999999999

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	11771870	6124532	9603344
Total Reads (QC-failed)	0	0	0
Duplicate Reads	0	0	0
Duplicate Reads (QC-failed)	0	0	0
Mapped Reads	11771870	6124532	9603344
Mapped Reads (QC-failed)	0	0	0
% Mapped Reads	100.0	100.0	100.0
Paired Reads	11771870	6124532	9603344
Paired Reads (QC-failed)	0	0	0
Read1	5885935	3062266	4801672
Read1 (QC-failed)	0	0	0
Read2	5885935	3062266	4801672
Read2 (QC-failed)	0	0	0
Properly Paired Reads	11771870	6124532	9603344
Properly Paired Reads (QC-failed)	0	0	0
% Properly Paired Reads	100.0	100.0	100.0
With itself	11771870	6124532	9603344
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	6611491	3365487	5299279
Distinct Fragments	5820494	3039596	4717080
Positions with Two Read	645243	277320	487950
NRF = Distinct/Total	0.88036	0.903167	0.890136
PBC1 = OneRead/Distinct	0.877232	0.901087	0.886985
PBC2 = OneRead/TwoRead	7.913175	9.87646	8.574602

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	70210	969
N1	44086	474
N2	34180	449
Np	78606	1037
N optimal	78606	1037
N conservative	70210	969
Optimal Set	pooled-pr1_vs_pooled-pr2	pooled-pr1_vs_pooled-pr2
Conservative Set	rep1_vs_rep2	rep1_vs_rep2
Rescue Ratio	1.1195841048283721	1.0701754385964912
Self Consistency Ratio	1.2898186073727327	1.0556792873051224
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	128631	106990

Top 300000 raw peaks from spp with FDR 0.01

Peak calling statistics

Peak region size

	rep1	rep2	idr_opt	overlap_opt
Min size	78.0	75.0	80.0	80.0
25 percentile	310.0	300.0	320.0	320.0
50 percentile (median)	310.0	300.0	320.0	320.0
75 percentile	310.0	300.0	320.0	320.0
Max size	318.0	357.0	369.0	369.0
Mean	309.9837441985214	299.9723712496495	314.2237222757956	319.9237971656108

Enrichment / Signal-to-noise ratio

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

	rep1	rep2
Number of Subsampled Reads	6257349	3183626
Estimated Fragment Length	205	210
Cross-correlation at Estimated Fragment Length	0.663186159585467	0.511843683508936
Phantom Peak	50	50
Cross-correlation at Phantom Peak	0.6633137	0.5113104
Argmin of Cross-correlation	1500	1500
Minimum of Cross-correlation	0.6596435	0.5073175
NSC (Normalized Strand Cross-correlation coeff.)	1.005371	1.008922
RSC (Relative Strand Cross-correlation coeff.)	0.9652533	1.133572

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	0.41402927497635234	0.3982027499954999
Synthetic AUC	0.4981449513076553	0.4974290742808691
X-intercept	0.019275978367159594	0.020321698696840886
Synthetic X-intercept	0.0	0.0
Elbow Point	0.524103454469257	0.5315771617873036
Synthetic Elbow Point	0.501278899506791	0.5044926676782259
JS Distance	0.029488110286022454	0.05425190579724528
Synthetic JS Distance	0.13881643508523692	0.15055758448436857
% Genome Enriched	45.34255323394999	45.171925203057334
Diff. Enrichment	7.917362077350276	10.613251498126658
CHANCE Divergence	0.06738027589916581	0.09046223797114067

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	0.5530817108921522	0.5147191654807257	0.44583206477270565	0.4625146868364799	0.4477207865395704	0.4622981804977099	0.5504159439422517	0.5515741598144521	0.5525604032095841

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	0.3349873343256371	0.22767801547247804	0.19919726111317568	0.3699099405567667

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	0.010788984288573758	0.005934910935985532	0.007039884843445997	0.011479570027539614

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