Quick start

This is quick start guide for impatient people.

Installation

Build and activate a conda environment and get XICRA repository:

## clone repo
git clone https://github.com/HCGB-IGTP/XICRA.git

## create conda environemt
conda env create -n XICRA -f XICRA_pip/devel/conda/environment.yml

## activate the environment
conda activate XICRA

## install latest python code
pip install XICRA

## install missing software
sh XICRA_pip/XICRA/config/software/installer.sh

Sometimes, the activation of the environment is done with source activate XICRA instead of conda activate XICRA. Check everything is fine by executing the config module:

XICRA config

Execute XICRA

The functionallity of XICRA is divided in modules. Each of them in charge of different parts of the analysis. Here we show how to run each of the modules:

  1. Prepation of the data: prep, QC, trimm, join.

    • XICRA prep --input input_folder --output output_folder: preparation of the data.

    • XICRA QC --input input_folder: quality analysis of the samples.

    • XICRA trimm --input input_folder --adapters_a XXXXX --adapters_A YYYYYYYY: trimming the adapters.

      • --adapters_a XXXXX: sequence of the 3’ adapter of R1

      • --adapters_A  YYYYYYYY: sequence of the 3’ adapters of R2.

    • XICRA join --input input_folder: join paired end reads.

  2. Quantification of RNA types: biotype

    • XICRA biotype --input input_folder --threads X --fasta file.fa --annotation file.gtf --limitRAM YYYY

      • --threads X: number of cores enabled to run the analysis.

      • --fasta file.fa: fasta file with the reference genome to map reads.

      • --annotation file.gtf: reference genome annotation in GTF format.

      • --limitRAM YYYY: given RAM in bytes to run the analysis. Note that: this process consumes high RAM values.

  3. miRNA analysis: miRNA

    • XICRA miRNA --input input_folder --threads X --software analyisis_tool

      • --threads X: number of cores enabled to run the analysis.

      • --software analyisis_tool: three options available, miraligner, optimir and sRNAbench.

Take into account that these are basic examples, each of the modules can be run with other different parameters. To see all the possible parameters of each module run -h. For example, for the trimm module:

XICRA trimm -h

Test example

Here we include a brief example on how to use XICRA.

First, we create a conda environment and install XICRA and its dependencies. See example details shown before. Then, we can test XICRA by using an example of 100 miRNA simulated and provided within the repository as an example of simulation.

## run XICRA example
ln -s ~/BMC_bioinformatics_paper/simulation/example/reads/

## prepare reads
XICRA prep --input reads/ --output_folder test_XICRA

## join reads
XICRA join --input test_XICRA --noTrim

## create miRNA analysis
XICRA miRNA --input test_XICRA --software miraligner optimir

## explore results
ls test_XICRA/report/

As a result, we will end up with a folder for each of the run analysis for every sample. Thus, in the miRNA folder, we will obtain the miRNA results for our samples, performed with two different softwares miraligner and optimir.

User data

In the presented example, nor QC, neither trimm steps were necessary (that is why --noTrim was added in the join command). However, with real data, running QC is highly recommended to check the quality of the samples (and filter outliers if necessary). Running the trimm command to eliminate the adapters will be necesary for NGS data.

On the other hand, the biotype was skipped as well. This step is only informative: it maps and annotates the reads and quantifies the RNA types present in each of the samples. Note that: this step is very time and RAM consuming.

The miRNA analysis can be performed whith any of the three available softwares miraligner, optimir and sRNAbench (they can be combined as seen in the example). Unfortunately, the donwloading of sRNAbench is no longer available, thus, only users with the software already installed will be able to run the miRNA analysis with it.

Finally, XICRA is also able to analyse single end reads, in this case --single_end should be added in the commands (and no join step would be necessary).

Deactivate environment

After finished the execution of any XICRA module or script, it is convenient to deactivate the environment. You can just close the terminal but it would be more appopiate to conveniently deactivate the environment first.

To do so, execute one of the following commands:

conda deactivate XICRA

source deactivate XICRA