npTranscript is a tool which was designed primarily for analysis of SARS-Cov-2 direct RNA sequence data. It works via analysis of BAM file of direct RNA sequencing reads aligned to a reference genome. It handles analysis of alignment CIGAR strings in order to cluster reads into different transcripts. It also enables analysis of 5'to 3' break points. Example of the output of this tool can be seen at https://doi.org/10.26188/5e86c8229aebb
npTranscript consists of two components - a java component, which utilises japsa ( https://github.com/mdcao/japsa) for analysis of CIGAR strings.
The second part consists of R scripts which generate visualisation of outputs of the java component
###INSTALLATION AND USAGE
To install java tool, change into the java subdirectory and run
bash install_mvn.sh
mvn clean package install
A precompiled jar is provided in the distribution.
The script run_example.sh provides an example of how you can run the java part of npTranscript.
The input required is
- 1 or more bam files (multiple bam files can be processed at the same time). We also include a script 'make_bams.sh' to recreate the minimap2 parameters and arguments used in our own analysis.
- a reference genome (we provide a reference coronavirus genome in data/SARS-Cov2/wuhan_coronavirus_australia.fasta.gz)
- a coordinate file (we provde one in data/SARS-Cov2/Coordinates.csv)
The output of the java program are the following files:
0readToCluster.txt.gz - master file of mapping of reads to clusters 0transcripts.txt.gz - transcript cluster file 0breakpoints.txt.gz.0.gz - file with break point information 0clusters.h5 - coverage information for each transcript.
These files can be further processed with the Rscripts provided
#RUNNING R SCRIPTS
Change into the resdir generated by the java program. run
Rscript npTranscript.R
npTranscript is currently maintained by Lachlan Coin (@lachlanjmc) and Dan Rawlinson.