seek for circular RNA in transcriptome (identifies deferentially expressed circRNAs between two samples)
Version: v1.0.0.b
Last Modified: 2019-04-25
Authors: Bioinformatics Lab, University of Louisville, Kentucky Biomedical Research Infrastructure Network (KBRIN)
circular RNA derived from exon 2 of Cdp gene, Rat, Hippocampus, Somata and Neuropil.
This circular RNA is highly expressed in Neuropil (31.9%) than Somata (11.9%).
1 Download seekCRIT
git clone https://github.com/UofLBioinformatics/seekCRIT.git
cd seekCRIT
2 Install required packages (Some prerequisite packages might require admin access rights. Please contact your system admin to install such packages.)
pip3 install -r Prerequisites.txt
3 Install seekCRIT
python setup.py install
4 testing seekCRIT with testrun.sh
In order to run testrun.sh:
./testrun.sh gtf/Rattus_norvegicus.Ensembl.rn6.r84.gtf fasta/rn6.fa
usage: seekCRIT.py [-h] -s1 S1 -s2 S2 -gtf GTF -o OUTDIR -t {SE,PE}
--genomeIndex GENOMEINDEX -fa FASTA -ref REFSEQ
[--threadNumber numThreads]
[--aligner aligner]
[--deltaPSI DELTAPSI] [--highConfidence HIGHCONFIDENCE]
[--libType {fr-unstranded,fr-firststrand,fr-secondstrand}]
[--keepTemp {Y,N}]
Identifying and Characterizing Differentially Spliced circular RNAs between
two samples
Required arguments:
===================
-s1 S1, --sample1 S1 fastq files for sample_1. Replicates are separated by
comma. Paired-end reads are separated by colon.
e.g.,s1-1.fastq,s1-2.fastq for single-end read. s1-1.R
1.fastq:s1-1.R2.fastq,s1-2.R1.fastq:s1-2.R2.fastq for
single-end read
-s2 S2, --sample2 S2 fastq files for sample_2. Replicates are separated by
comma. Paired-end reads are separated by colon.
e.g.,s2-1.fastq,s2-2.fastq for single-end read. s2-1.R
1.fastq:s2-1.R2.fastq,s2-2.R1.fastq:s2-2.R2.fastq for
single-end read
-gtf GTF, --gtf GTF The gtf annotation file. e.g., hg38.gtf
-o OUTDIR, --output OUTDIR
Output directory
-t {SE,PE}, --readType {SE,PE}
Read type. SE for Single-end read, PE for Paired-end read
--genomeIndex GENOMEINDEX
Genome indexes for the aligner
-fa FASTA, --fasta FASTA
Genome sequence. e.g., hg38.fa
-ref REFSEQ, --refseq REFSEQ
Transcriptome in refseq format. e.g., hg38.ref.txt
optional arguments:
====================
-h, --help show this help message and exit
--threadNumber numberOfThreadsk
Number of threads for multi-threading feature [default = 4]
--aligner aligner aligner to use(for now it supports only STAR but we are working on it to support more aligners)
--deltaPSI DELTAPSI Delta PSI cutoff. i.e., significant event must show
bigger deltaPSI than this cutoff [default = 0.05]
--highConfidence HIGHCONFIDENCE
Minimum number of circular junction counts required [default = 1]
--libType {fr-unstranded,fr-firststrand,fr-secondstrand}
library type used by Tophat aligner [default ='fr-unstranded']
--keepTemp {Y,N} Keep temp files or not [default='Y']
python3 seekCRIT.py -o PEtest -t PE -fa fa/hg19.fa -ref ref/hg19.ref.txt --genomeIndex /media/bio/data/STARIndex/hg19 -s1 testData/231ESRP.25K.rep-1.R1.fastq:testData/231ESRP.25K.rep-1.R2.fastq,testData/231ESRP.25K.rep-2.R1.fastq:testData/231ESRP.25K.rep-2.R2.fastq -s2 testData/231EV.25K.rep-1.R1.fastq:testData/231EV.25K.rep-1.R2.fastq,testData/231EV.25K.rep-2.R1.fastq:testData/231EV.25K.rep-2.R2.fastq -gtf testData/test.gtf --threadNumber 12
python3 seekCRIT.py -o SEtest -t SE -fa fa/hg19.fa -ref ref/hg19.ref.txt --genomeIndex /media/bio/data/STARIndex/hg19 -s1 testData/231ESRP.25K.rep-1.R1.fastq,testData/231ESRP.25K.rep-1.R2.fastq,testData/231ESRP.25K.rep-2.R1.fastq,testData/231ESRP.25K.rep-2.R2.fastq -s2 testData/231EV.25K.rep-1.R1.fastq,testData/231EV.25K.rep-1.R2.fastq,testData/231EV.25K.rep-2.R1.fastq,testData/231EV.25K.rep-2.R2.fastq -gtf testData/test.gtf --threadNumber 12
Field | Description |
---|---|
geneName | Name of gene |
isoform_name | name of isoform |
chrom | chromosme |
strand | strand (+/-) |
txStart | Transcription start position |
txEnd | Transcription end position |
cdsStart | Coding region end |
exonCount | Number of exons |
exonStarts | Exon start positions |
exonEnds | Exon end positions |
See details in the example file
Field | Description |
---|---|
chrom | chromosome |
circRNA_start | circular RNA 5' end position |
circRNA_end | circular RNA 3' end position |
strand | DNA strand (+/-) |
exonCount | number of exons included in the circular RNA transcript |
exonSizes | size of exons included in the circular RNA transcript |
exonOffsets | offsets of exons included in the circular RNA transcript |
circType | circRNA, ciRNA, ccRNA |
geneName | name of gene |
isoformName | name of isoform |
exonIndexOrIntronIndex | Index (start from 1) of exon (for circRNA) or intron (for ciRNA) in given isoform |
FlankingIntrons | Left intron/Right intron |
CircularJunctionCount_Sample_1 | read count of the circular junction in sample # 1 |
LinearJunctionCount_Sample_1 | read count of the linear junction in sample # 1 |
CircularJunctionCount_Sample_2 | read count of the circular junction in sample # 2 |
LinearJunctionCount_Sample_2 | read count of the linear junction in sample # 2 |
PBI_Sample_1 | Percent Backsplicing Index for sample # 1 |
PBI_Sample_2 | Percent Backsplicing Index for sample # 2 |
deltaPBI(PBI_1-PBI_2) | difference between PBI values of two samples |
pValue | pValue |
FDR | FDR |
To calculate the significane of differentially expressed circular RNAs, use the criteria:
At least 5% changes in percent back-spliced in (PBI) or |deltaPBI|>=5%
FDR < 0.05
Copyright (C) 2017 . See the LICENSE file for license rights and limitations (MIT).