A tool to simulate RNA-seq reads from circular and linear RNAs
Clone the GIT repository, enter the repo and run
./utils/install.sh
You will need a reference genome/chromosomes and gene annotation, for instance
mkdir -p annotation
cd annotation
wget ftp://ftp.ensembl.org/pub/release-95/fasta/homo_sapiens/dna/Homo_sapiens.GRCh38.dna.chromosome.1.fa.gz
zcat Homo_sapiens.GRCh38.dna.chromosome.1.fa.gz | sed "s/^>1/>chr1/" > chr1.fa
wget ftp://ftp.ebi.ac.uk/pub/databases/gencode/Gencode_human/release_29/gencode.v29.annotation.gtf.gz
zgrep -w "^chr1" gencode.v29.annotation.gtf.gz > chr1.gencode.v29.annotation.gtf
Then, make your project directory and set the simulation parameters in a Python file named vars.py, such that in the test directory.
Now, you are ready to generate the simulated reads by calling
/path/to/ccp_sim/ccp_sim.sh
GENE_ANNO: Input GTF formatted annotation file name
default: annotation/gencode.v29.annotation.gtf
REFSEQ_DIR: Directory of reference sequence(s)
default: annotation/
CIRI_SIM_OPT: Options for the CIRI_simulator.pl script
default: -C 20 -LC 0 -R 1 -LR 1 -L 101 -E 1 -CHR1 1 -M 250 -M2 450 -PM 0 -S 70 -S2 0 -SE 0 -PSI 10
Simulated reads will be in the ccp_reads directory.
Simulated circular RNAs with read counts are saved in the cirisim_pred/cirias_tp.csv table.
Processed annotation is saved in the ccp_anno/annotation4simulations.gtf file.
Summary counts and annotation status for each annotated transcript are saved in the ccp_anno/summary.txt file. Annotation cases refer to the table below:
| case | circRNA expression | linear trx expression | linear trx annotation | meaning |
|---|---|---|---|---|
| 1 | yes | yes | yes | circRNA from expressed known trx |
| 2 | yes | yes | no | circRNA from novel trx |
| 3 | yes | no | yes | only circRNA from known trx |
| 4 | yes | no | no | novel circRNA gene |
| 5 | no | yes | yes | just no circRNA |
| 6 | no | yes | no | only novel trx |
| 7 | no | no | yes | known trx not expressed |
| 8 | no | no | no | nothing expressed, nothing known |