<h1>VDJ Puzzle</h1> <h3>A new version of VDJPuzzle is available at</h3> <a href="https://bitbucket.org/kirbyvisp/vdjpuzzle2">https://bitbucket.org/kirbyvisp/vdjpuzzle2</a><br><br><br>

VDJ Puzzle [<a href="#Ref1">1</a>] is a method to build T-Cell Receptor and B-Cell Receptor assembly from single cell RNA sequencing data.

<h2>Requiraments</h2> VDJ Puzzle requires the following tools: <ul> <li><a href="">TopHat2</a> for alignament;</li> <li><a href="">Trinity 2.0.6</a> for denovo assembly of TCR;</li> <li><a href="">Ensembl GRCh37</a> as refernce genome, but it can be easily adapted to other reference genomes (<a href="#generateBED">here</a> it is explained how);</li> <li><a href="https://github.com/mikessh/migmap">MiGMAP v0.9.7</a> to build a detailed information table (incoroporates IgBlast);</li> <li><a href="">Java 1.7</a> is required by Trinity 2.0.6;</li> </ul> <h2>Installation</h2> 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 ``` <h2>Usage</h2>

./run_tcr.sh directory_name [options]

<h2>Additional Information</h2>

<a name="generateBED"></a><h3>How to generate new BED files for different reference genomes</h3> 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 <i>chromosome</i>{tab}<i>starting position</i>{tab}<i>ending position</i> for each TRA gene and rename the file as "TRA.bed". Repeat for TRB adding the command

grep "TRBD" genes.gtf > TRBD.csv

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

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

<h2>Example</h2> We provide some example files to test VDJPuzzle. These sequences are a subset of two cells part of the dataset used in [<a href="#Ref1">1</a>] (Link to dataset cooming soon).<br> from VDJPuzzle directory ```bash ./VDJPuzzle.sh Example ``` <h2>Further Questions?</h2> You can post any question or issue to our <a href="https://groups.google.com/forum/#!forum/vdj-puzzle">Google group</a> <br> or by email ( s.rizzetto at student.unsw.edu.au ) <h2>Cite us</h2> [<a name="Ref1">1</a>] 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

VDJPuzzle

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README