Pypette is a collection of command line utilities and libraries for analyzing biological data. Current functionality includes:

• Gene expression quantification
• Copy number analysis
• Mutation and SNP analysis
• Chromosomal rearrangement analysis
• FASTA, VCF and SAM file manipulation
• Other miscellaneous functionality

For performance reasons, it is recommended that the software is run using PyPy, a high performance implementation of the Python language. You can build PyPy from source, or download portable binaries.

To install Pypette, download the latest release and extract it to a folder. Then run the Makefile and add the subfolder bin/ to your PATH. See below for an example:

wget --content-disposition https://github.com/annalam/pypette/archive/0.7.1.tar.gz
tar -xzf pypette-0.7.1.tar.gz
cd pypette-0.7.1
make
export PATH=/some/folder/pypette-0.7.1/bin:$PATH

Some Pypette functionality requires external software to be installed:

• samtools: required for mutation and structural variant calling
• Bowtie: required for structural variant calling
• ANNOVAR: required for mutation annotation

Download and prepare a reference genome for use in split read analysis:

wget ftp://ftp.ccb.jhu.edu/pub/data/bowtie_indexes/hg19.ebwt.zip
unzip hg19.ebwt.zip
fasta flatten <(bowtie-inspect hg19) hg19

Identify structural variants based on read pairs and split reads. Split reads must overlap at least 25bp on both sides of the breakpoint:

breakfast detect -a25 test.bam hg19 test
breakfast detect -a25 control.bam hg19 control

Construct a blacklist of false positive regions based on the control sample:

breakfast blacklist control.sv > blacklist.txt

Only keep structural variants with at least one read pair and three split reads of evidence, or at least 10 split reads (one -r option must be satisfied). Also discard blacklisted structural variants found in the control:

breakfast filter -r 1-3-0 -r 0-10-0 --blacklist=blacklist.txt test.sv > filtered.sv

Annotate structural variants with information about nearby genes:

wget ftp://ftp.ensembl.org/pub/release-74/gtf/homo_sapiens/Homo_sapiens.GRCh37.74.gtf.gz
gtf to gene bed Homo_sapiens.GRCh37.74.gtf.gz > ensembl_genes.bed
breakfast annotate -b ensembl_genes.bed filtered.sv > annotated.sv