Overview

Here, we describe the RNA-RNA Interaction Scan (rriScan), a universal software for analyzing RNA-RNA interactome sequencing data, such as MARIO, PARIS, PARIS2 and LIGR-seq data.

• System requirements
• Run time
• Installation
• Input
• Output
• Basic usage
• Run rriScan on testing dataset
• Acknowledgements
• Contact

System requirements

The software package was tested on Linux systems with the following specifications:

• RAM: 64GB
• CPU: 16+ cores

Run time

Written in C and C++, rriScan is highly efficient for analyzing RNA-RNA interactome sequencing data. After testing, most tasks are completed within 10 minutes.

Installation

• Installing rriScan on a Linux server is straightforward. Use the following commands:

  #Assume your installation directory is /username/software
  
  cd /username/software
  
  git clone https://github.com/kerenzhou062/rriScan.git
  
  cd ./rriScan
  
  sh install.sh
  
  export PATH=$PATH:/username/software/rriScan/bin

Input

rriScan requires the following input files:

• Genome sequence in FASTA format: Specify using the --fa argument. Chromosome names in this file must match those in the input bam file.

• FAI index file: Specify using the --fai argument. Generate this file using samtools:

  #Example: Generate an FAI index for hg38.fa  
  
  samtools faidx hg38.fa

• BAM file: Specify using the --bam argument. This file should contain sequence alignment data generated by STAR software and stored as Aligned.sortedByCoord.out.bam.

• Junction file: Specify using the --jun argument. This file contains junction information, which can be generated by STAR software with the --chimOutType Junctions parameter and stored as Chimeric.out.junction.

• FASTQ or FASTA file: An optional file containing sequencing reads.

Output

Here’s the description of the columns in the output files:

Column name	Description
lChrom	Chromosome name of left pair
lChromStart	Start coordinate of left pair (0-base)
lChromEnd	End coordinate of left pair
lName	Name of left pair
lScore	The quality of the alignment from left paired RNA
lStrand	Strand of left pair
rChrom	Chromosome name of right pair
rChromStart	Start coordinate of right pair (0-base)
rChromEnd	End coordinate of right pair
rName	Name of right pair
rScore	The quality of the alignment from right paired RNA
rStrand	Strand of right pair
lociNum	Chromosome name
gapDist	Gap distance between pairs if they are in the same chromosome
readSeq	Sequencing reads
chimericSeq	Full sequence of the chimeric
chimericStruct	Predict structure of chimeric
MFE	Minimum free energy
rriType	Type of RNA-RNA interaction
lAlignSeq	Aligned sequence of left pair
pairs	Base pairings
rAlignSeq	Aligned sequence of right pair
pairNum	The maximum continuous perfect pairings
alignScore	The quality of the alignment between left paired and right paired RNA
loReadNum	Read number of left pair
roReadNum	Read number of right pair

Basic Usage

The available options for rriScan are as follows:

Usage:  rriScan [options] --fa <fasta file> --fai <fai file> --bam <mapped alignments> --jun <junctions>
File format for mapped alignments is BAM
[options]
-v/--verbose                   : verbose information
-V/--version                   : rriScan version
-h/--help                      : help informations
-S/--small                     : small genome
--fa                           : genome FASTA file. [required]
--fai                          : genome fai file, an index file for fasta file. [required]
--bam                          : alignment file, BAM format. [required]
--jun                          : junction file from STAR software, junction format. [required]
--read                         : read file[fastq or fasta]. [optional]
-o/--output <string>           : output file
-l/--min-seg-len <int>         : minimum length of segments in a chimera [default>=15]
-m/--min-read-number <int>     : minimum read number for chimera [default>=1]
-M/--max-mfe <double>          : maximum MFE in duplex[default<=-5.0]
-p/--min-pair <int>            : minimum pair number in duplex [default>=0]
-s/--min-score <int>           : minimum alignment score in duplex [default>=5]
-g/--min-gap <int>             : minimum gaps between two segments [default>=1]

Run rriScan on testing dataset

Please check this guide to learn how to run rriScan on a testing dataset.

Acknowledgements

Thanks to everyone who contributed to the public codes and libraries (e.g., BamTools) used by rriScan.

Contact

• Jian-Hua Yang yangjh7@mail.sysu.edu.cn, RNA Information Center, School of Life Sciences, Sun Yat-Sen University
  
• Keren Zhou kzhou@stjude.org, Department of Pathology, St. Jude Children’s Research Hospital, Memphis, TN, USA