bcmap

Maps barcodes to a reference genome and returns genomic windows from which the barcoded reads most likely originate. Each window is assessed with a quality score representing the trustworthines of the mapping. A barcode index is constructed alongside the mapping and can be used to quickly retrieve all reads belonging to a barcode. For ease of use we provide a snakemake workflow to extract all reads from user defined regions of interest.

Prerequisites

• gcc version 7.2.0

Installation

git clone https://github.com/kehrlab/bcmap.git
cd bcmap
make

Data requirements

• Paired-end Linked-reads

• Barcodes are stored in BX:Z: flag of read Ids

• Sorted by barcode (use i.e. bcctools (only for 10x genomics linked-reads) or samtools )

To trimm, correct and sort barcodes with bcctools use the following command in the bcctools folder:

./script/run_bcctools -f fastq first.fq.gz second.fq.gz

Commands

For detailed information on Arguments and Options:

./bcmap [command] --help

index

Builds an minimized open addressing k-mer index of the reference genome. The index is required to run "map".

./bcmap index reference.fa [options]

map

Maps the barcodes of the provided readfiles to the reference and creates a barcode index of the readfiles to quickly retrieve all reads of a given barcode.

./bcmap map readfile1.fastq readfile2.fastq [options]

Content of output bed-file:

• chromosome startposition endposition barcode mapping_score

Bcmap returns a output.hist file that can be ploted using plot_score_histogram.py resulting in a plot like the one above. To create a set of mappings with very high precision (at the cost of some recall), the local minimum inbetween the two peaks should be set as the score threshold. A lower theshold yields better recall at the cost of precision, a higher threshold is not recomended.

get

Returns all reads of the given barcodes. Barcodes can be provided directly as argument or in a file.

./bcmap get readfile1.fastq readfile2.fastq Barcodes [options]

Example

This small example demonstrates how to use bcmap and allows you to check if it is properly installed. Navigate to the bcmap folder and run the commands listed below.

# building the index for chr21.fa
./bcmap index example/chr21.fa -o example/Index
# mapping the reads of readfile 1 and 2 to chromosome 21
./bcmap map example/readfile.1.fq example/readfile.2.fq -i example/Index -r example/ReadIndex -o example/results.bed
# extracting the first barcode from the results
awk '{if(NR==1) print($4)}' example/results.bed > example/FirstBarcode.txt
# extracting all reads belonging to the first barcode
./bcmap get example/readfile.1.fq example/readfile.2.fq example/FirstBarcode.txt -r example/ReadIndex -o example/readsOfFirstBarcode
# extracting reads of barcode AACATCGCAAACAGTA
./bcmap get example/readfile.1.fq example/readfile.2.fq AACATCGCAAACAGTA -r example/ReadIndex -o example/readsOfAACATCGCAAACAGTA