T cell Receptor and B cell Receptor assembly from single cell RNA sequencing data
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BED_files
Example
java1.8
Method2.sh
Method2_part3.sh
README.md
VDJPuzzle.sh
run_tcr_method2_sc_1file.sh
run_tcr_method2_sc_1file_part3.sh
run_tcr_method2_sc_part2.sh

README.md

VDJ Puzzle

A new version of VDJPuzzle is available at

https://bitbucket.org/kirbyvisp/vdjpuzzle2


VDJ Puzzle [1] is a method to build T-Cell Receptor and B-Cell Receptor assembly from single cell RNA sequencing data.

Requiraments

VDJ Puzzle requires the following tools:
  • TopHat2 for alignament;
  • Trinity 2.0.6 for denovo assembly of TCR;
  • Ensembl GRCh37 as refernce genome, but it can be easily adapted to other reference genomes (here it is explained how);
  • MiGMAP v0.9.7 to build a detailed information table (incoroporates IgBlast);
  • Java 1.7 is required by Trinity 2.0.6;

Installation

To install VDJ Puzzle is sufficient to unzip it in a folder and update the paths to the required softwares. You can change the paths in the "#CONFIGURE SCRIPT PATHS" section located in VDJPuzzle.sh. ```bash export MIGMAP=/path_to_migmap/migmap(version).jar ``` The remaining tools are expected to in the /usr/bin/ directory if not you can change their path. ```bash export trinitypath=Trinity export TOPHAT=tophat2 export BOWTIE=bowtie2 ``` in the next section #CONFIGURE REFERENCE PATHS you need to update the link to Ensembl reference genome ```bash export MIGMAP=/path_to_migmap/migmap(version).jar ```

Usage

./run_tcr.sh directory_name [options]

Additional Information

How to generate new BED files for different reference genomes

If you want to use a different reference genome you need to generate new BED files containing the position of the constant segment transcripts.

grep "TRAC" genes.gtf > TRAC.csv
grep "TRAV" genes.gtf > TRAV.csv
grep "TRAJ" genes.gtf > TRAJ.csv
cat TRAC.csv TRAV.csv TRAJ.csv > TRA.csv

edit TRA.csv in order to have a table with chromosome{tab}starting position{tab}ending position for each TRA gene and rename the file as "TRA.bed". Repeat for TRB adding the command

grep "TRBD" genes.gtf > TRBD.csv

Finally, you need to update the position of the new BED files in run_sc.sh

TCRA=/path_to_bed_files/TRA.bed
TCRB=/path_to_bed_files/TRB.bed

Example

We provide some example files to test VDJPuzzle. These sequences are a subset of two cells part of the dataset used in [1] (Link to dataset cooming soon).
from VDJPuzzle directory ```bash ./VDJPuzzle.sh Example ```

Further Questions?

You can post any question or issue to our Google group
or by email ( s.rizzetto at student.unsw.edu.au )

Cite us

[1] Auda Eltahla\*, Simone Rizzetto\*, Mehdi Rasoli\*, Brigid Betz-Stablein, Vanessa Venturi, Katherine Kedzierska, Andrew R Lloyd, Rowena A Bull and Fabio Luciani. Linking the T cell receptor to the single cell transcriptome in antigen-specific human T cells. Immunology and cell biology, 2016, doi: 10.1038/icb.2016.16G