/siRNA-mapped-to-Niben-length-distribution

Primary LanguageR

This pipeline describes the collection,calculation and plotting of siRNA length distribution that map to the Nicotiana benthamiana transcriptome. The Nicotiana benthamiana v2.6.1 transcriptome was downloaded from solgenomics.net

Citation

When using this software cite the following:
(https://github.com/olgatsiouri1996/siRNA-mapped-to-Niben-length-distribution. Kellari et al., unpublished)

Publication

When using this software cite the following publication:
Kellari L., Dalakouras A., Tsiouri O., Vletsos P., Katsaouni A., Uslu V. V. and Papadopoulou K. K. Cross-kingdom RNAi induced by a beneficial endophytic fungus to its host requires transitivity and amplification of silencing signals (unpublished)

Dependences/Installation

  1. docker
  2. docker container used to retrieve the 21-28nt siRNA counts per samples:
docker pull olgatsiouri/sinra_21_28nt_length_calculator_all
  1. docker container used to plot the data:
docker pull pegi3s/r_data-analysis

Usage

The sinra_21_28nt_length_calculator_all performs the following analysis:
1). uses bowtie-build to build indexes of user specified fasta file
2). runs bowtie -v 0 --norc --best --strata -a per each sample(samplenames specified in list)
3). selects siRNAs with length 21-28 nt in 8 files per length with cutadapt for each sample
4). uses grep for each of the 21nt to 28nt to calculate the number of reads per sample

The final result is 8 .txt files with a num prefix, 1 for each length, where each line is each sample in list To run the whole pipeline run the following:

  1. run the sinra_21_28nt_length_calculator_all container:
docker run -v /media/linuxubuntu2004/INTENSO/loukia/cleaned/dock/niben:/database -v /media/linuxubuntu2004/INTENSO/loukia/cleaned/dock:/input -v /media/linuxubuntu2004/INTENSO/loukia/cleaned/dock:/output -v /media/linuxubuntu2004/INTENSO/loukia/cleaned/dock:/list olgatsiouri/sinra_21_28nt_length_calculator_all niben 4

The format for linux and macos is:

docker run -v /path/to/database:/database -v /path/to/input:/input -v /path/to/output:/output -v /path/to/list:/list olgatsiouri/sinra_21_28nt_length_calculator_all database_name thread_count

The format for windows is:

docker run -v C:\path\to\database:/database -v C:\path\to\input:/input -v C:\path\to\output:/output -v C:\path\to\list:/list olgatsiouri/sinra_21_28nt_length_calculator_all database_name thread_count

where /path/to/database is the path to the folder containing the Nicotiana benthamiana transcriptome, /path/to/input is the path to the folder containing trimmed fastq.gz files to be mapped and database_name is the name of the indexed transcriptome files

  1. Put the groups.txt and .R files on the same folder as the rest of the num*.txt files
  2. Run the following to plot the results(in this example all txt files and the .R script are in the /home/linuxubuntu2004/Desktop folder):
docker run --rm -it -v /home/linuxubuntu2004/Desktop:/data pegi3s/r_data-analysis Rscript /data/length_distribution_mapped_to_niben_transcripts_calculation.R

the group.txt file(that in this case contains 4 different conditions with 2 technical replicates each) can be modified for use in other projects