Data processing steps

circRNA identification

1. Map RNAseq to genome using bwa. The mapped sam will be used for circRNA calling.

   • bwa mem -T 19 GRCh38.d1.vd1.fa RNAseq_R1.fastq RNAseq_R2.fastq -o bwa-pe-for-CIRI.sam

2. circRNA calling using CIRI.

   • perl CIRI2.pl 16 -I bwa-pe-for-CIRI.sam -O results_CIRI.txt -F GRCh38.d1.vd1.fa -A gencode.v34.basic.annotation.gtf

Prepare index for linear and circRNA quantification

1. Add gene names to CIRI outputs by the inhouse script.

   • perl 1_add_gene_name_to_CIRI_results.pl

2. Add circRNA to gene annotation in the gft format by the inhouse script.

   • perl 2_add_linear_circular_isoform_to_gtf.pl

3. Extract both linear and circRNA transcripts using RSEM.

   • rsem-extract-reference-transcripts GRCh38.d1.vd1.linear.and.circrna.isoforms 0 gencode.v34.basic.annotation.gtf None 0 genome/GRCh38.d1.vd1.fa

4. Add gene names to CIRI outputs by the inhouse script.

   • perl 1_add_gene_name_to_CIRI_results.pl

Run RSEM to quantifiy linear and circRNA isoforms

Summarize RSEM output