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typing_instruction.md

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Vibrio cholerae typing from Illumina data

This is a guide for taking reads generated on an Illumina sequencer and mapping them against specific genes (ctxA, wbeO1, wbfO139, tcpA classical, tcpA El Tor, toxR) for the purpose of determining sample serotype.

Setting up your working directory

This script assumpes the following:

  • That you have already set up your working directory as described in the assembly instructions. Make sure you know where the raw fastq files are stored and make sure they follow the naming convention

  • That the scripts folder inside your working directory contains the map_genes.sh script.

  • That all six gene-specific reference FASTAs are contained in a directory called typing. If you cloned this git repository, these reference files can be found in ref_genomes/typing.

Once you have the required files in your working directory, navigate to this directory and create a new folder where you will store your typing results:

cd my-project

mkdir typing_results

Running the typing script

Run the typing script as follows:

bash scripts/map_genes.sh sample-name my-project path-to-raw-data num-lanes num-cores ref-dir output-dir

Where sample-name is the 'SampleName' prior to '_L00...' in your fastq files,my-project is the absolute path to your project directory, and path-to-raw-data is the path to your raw fastq files. num-lanes is the number of lanes you used to run this sample. num-cores should be less or equal to the number of processors on your computer.ref-dir should be the directory that contains the gene-specific FASTA files (usually my-project/ref_genomes/typing). output-dir will be the folder to store typing results (my-project/typing_results).

Evaluating typing results

The typing script will produce one output file (samplename_typing_summary.txt). This file will look something like this:

SAMPLENAME      gene    reads_mapped    reads   frac_mapped     mean_depth      pos_covered     total_pos       frac_covered
ERR3039943      ctxA    96      588182  .0001632        27.4582 777     777     1.0000000
ERR3039943      tcpA_classical  87      588182  .0001479        9.74046 1048    1048    1.0000000
ERR3039943      tcpA_eltor      96      588182  .0001632        28.3126 675     675     1.0000000
ERR3039943      toxR    134     588182  .0002278        30.8923 891     891     1.0000000
ERR3039943      wbeO1   3660    588185  .0062225        38.4333 21122   21122   1.0000000
ERR3039943      wbfO139 1923    588187  .0032693        9.17084 46721   46721   1.0000000

These files may be easier to view in a spreadsheet software such as Excel.