XCALIBR

Introduction

xcalibr - eXtracting Counting and LInking to Barcode References

xcalibr processes short reads and extracts a table of counts matched to index sequences and tag sequences

Method

Before the short reads are counted they are first processed into a binary structure (hash), this ensures only unique reads processed which speeds up the process.

xcalibr uses a template sequence to locate and extract the barcodes. Only exact matches to the reference sequence are currently supported. The reference sequences should be provided as fasta files. If no reference is present all barcodes at the requested location are reported which will result in large output files. You can use xcalibr reduce to reduce remove barcodes with low counts. xcalibr supports three

Dependencies

The program is implemented in perl and currently uses only core modules.

Command line tools

A wrapper bash script can be used to call the different tools in the package.

xcalibr hash
xcalibr analyze
xcalibr extract
xcalibr reduce

hash

Run hash on a FastQ file. This file may be gzipped. Hash also reads (uncompressed FastQ) from STDIN if no filename is provided.

analyze

Use analyzehash to debug the input data. It will print a summary table given de hash file and a range. You can use is to check constant parts or get a simple overview when relatively few barcodes have been sequenced.

extract

The workhorse program. Provide a template that xcalibr uses to locate and extract the barcodes. CGAT are used to encode constant sequences and X, Y and Z be be used as placeholder for the barcodes to be extracted. If only 1 or 2 placeholders are used a single table is generated. I three placeholders are used one is used to generate multiple output files.

reduce

Use 'xcalibr reduce' to filter the output table. Use the --mincount argument to specify the minimum number of reads that must be cocounted for a barcode to pass.

Examples

If the xcalibr path is in your PATH you can use the following example:

cat testdata.fq | xcalibr hash testdata.bin
xcalibr analyze testdata.bin 7-22
xcalibr extract --template X6CAGGCGCTTAGGATCCY15  testdata.bin > res-all.txt
xcalibr extract --template X6CAGGCGCTTAGGATCCY15  testdata.bin | xcalibr reduce --mincount 5 > res-min5.txt

You can use zcat if your FastQ file is gzipped.