RNA-seq using a reference genome

An integrated worklow based on genome-mapping and DE gene assessment to conduct RNAseq data analyses in Colosse

WARNING

The software is provided "as is", without warranty of any kind, express or implied, including but not limited to the warranties of merchantability, fitness for a particular purpose and noninfringement. In no event shall the authors or copyright holders be liable for any claim, damages or other liability, whether in an action of contract, tort or otherwise, arising from, out of or in connection with the software or the use or other dealings in the software.

Documentation

Note that ALL the scripts must be launched from the root folder rna-seq_mapping_workflow/

1. Clone git hub directory

git clone https://github.com/jleluyer/rna-seq_mapping_workflow

2. Prepare utilities

cd rna-seq_mapping_workflow

./00_scripts/00_import_trinity.sh

This script will create two directories 00_scripts/trinity_utils and 00_scripts/trinotate_utils

cd 00_scripts/trinity_utils/
make
make plugins
cd ../..

still have to properly configurate trinity on Colosse

3. Import data

cp /path/to/the/data/repository/*.gz 02_data

4. Trimming the data

• Import univec.fasta

wget 01_info_files/univec.fasta ftp://ftp.ncbi.nlm.nih.gov/pub/UniVec/UniVec

Add your specific adaptors if absent in the database.

• Trimming

Two scripts are provided for Single-End or Paired-end data, 00_scripts/01_trimmomatic_se.sh and 00_scripts/01_trimmomatic_pe.sh, respectively.

First you need to change the userID and userEmail for your proper info in ./00_scripts/colosse-jobs/01_trimmomatic_pe.sh

Then launch:

./00_scripts/colosse-jobs/01_trimmomatic_pe.sh

You may also want to check the quality of your data prior to trimming using 00_scripts/utility_scripts/fastq.sh. This will require to have fastQC installed in your $PATH.

5. Index reference genome with GMAP

msub /00_scripts/02_gmap_index.sh

6. Mapping

Before launching the mapping, make sure you changed the userID and userEmail information in the script ./00_scripts/colosse_jobs/03_gmap_mapping_jobs.sh

Then launch:

./00_scripts/colosse_jobs/03_gmap_mapping_jobs.sh 

This script will create a job by individual.

7. Assess reads count

Before launching the mapping, make sure you changed the userID and userEmail information in the script ./00_scripts/04_htseq_count.sh

msub /00_scripts/04_htseq_count.sh

8. Downstream analysis

Notes

Dependencies

Softwares

GSNAP

HTseq

Bowtie

java 1.7 or higher

R

Citations

Licence