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Introduction

CIMS analysis is a computational method to analyze HITS-CLIP data for determining the exact protein-RNA crosslink sites and thereby mapping protein-RNA interactions at the single-nucleotide resolution. This method is based on the observation that UV cross linked amino-acid-RNA adduct introduce reverse transcription errors in cDNAs at certain frequencies, which are captured by sequencing and comparison of CLIP tags with the reference genome. More details can be found in the following reference:

Zhang, C. †, Darnell, R.B. † 2011. Mapping in vivo protein-RNA interactions at single-nucleotide resolution from HITS-CLIP data. Nat. Biotech. 29:607-614.

This brief document provides the most critical information about the algorithm, and is expected to be expanded into a more detailed tutorials in the near future.

Download

Source code:

• czplib (perl): a perl library with various functions for genomic/bioinformatic analysis. (download from SourceForge.net)
• CIMS (perl): the core algorithm. (download from SourceForge.net)

Installation

Prerequisites

This software is implemented with perl . It also relies on several standard linux/unix tools such as grep, cat, sort, etc. We have tested the software on RedHat Linux, although it is expected to work on most unix-like systems, including Mac OS X.

Steps to install the software

• Download the perl library files czplib, if not already.

Decompress it and move it to a place you like

$tar zxvf czplib.v1.0.x.tgz
$mv czplib /usr/local/lib

Add the library path to the environment variable, so perl can find it.

PERL5LIB=/usr/local/lib/czplib

• Download CIMS codes, if not already.

Decompress it and move it to a place you like

$tar zxvf CIMS.v1.0.x.tgz
$cd CIMS
$chmod 755 *.pl
$mv CIMS /usr/local/CIMS

Add the dir to your $PATH environment variable.

Get started

The key script one needs to run is CIMS.pl, which will take two BED files as input: a list of unique CLIP tags (properly mapped to the reference genome), and the coordinates of mutations (deletions, insertions, or substitutions) in the reference genome and relative the CLIP tags. It is critical to make sure that

• analyze one type of mutations at a time.
• the 4th column of the mutation BED file should match the name of the CLIP tag in the first BED file.
• the coordinates of mutations relative to the CLIP tag (from the 5' end of the Watson strand, 0-based) is correctly specified in the 5' column of the second BED file.
• only mutations in unique CLIP tags should be included.

Now you can run something like

perl /usr/local/bin/CIMS/CIMS.pl -v -n 5 -p -c ./cache_del  test.uniq.bed test.uniq.del.bed test.uniq.del.CIMS.txt