A software to design PCR primers for a a subset of sequences which belong to a greater set
Genprimers is a software aimed to produce PCR primers. Unlike other primers programs, such as primer3, it is focused on generating specific primers for a set the sequences (defined as targets) which are a subset of a greater set of sequences (defined as the universe). This is very useful for example when we have 16S amplicon experiments from ambiental samples, which have a lot of different miroorganisms, and we are interested in detecting spefific phylums or clades. Genprimers is also useful when we have to amplify a specific biomarker of a subset of individuals belonging to a greater set of closely- related microorganism.
- Bowtie 1
- Python 2.7
For the moment, Bowtie 1 is necessary to run genprimers. In the future, this won’t be necessary.
How to install
Genprimers is available in the Python pip repository
$ pip install genprimers
Clone the git repository in your local machine
$ git clone email@example.com:lbmg/genprimers.git
Enter to the cloned repository
$ cd genprimers
Install using the python install framework
$ python setup.py install
Generating the index for the universe
In order to run genprimers, the input FASTA file must be indexed with Bowtie (for the moment)
$ bowtie-build universe.fna universe.fna
Assuming we already have the FASTA file of the universe indexed with bowtie and a one-column file containing the IDs of a subset of sequences of the universe we want to amplify (targets_ids.txt in this example), run genprimers is as easy as:
$ genprimers primers universe.fna targets_ids.txt output_folder
The list of targets IDs can be passed through the standard input
$ cat targets_ids.txt | genprimers primers universe.fna output_folder
Listing all the sequences in the Universe
In order to run Genprimers you need the IDs of the targets in the FASTA file of the universe, but often we don’t know those identifier. To list the IDs in the universe, and their respectives descriptions, we can use the list command from genprimers.
$ genprimers list unvierse.fna
We can filter the output list to report only those sequences belonging to some class and then use that list to design primers. In the example below we list all the sequences that fit the Gluconobacter genus description and produce new primers for this subset:
$genprimers list universe.fna | grep Gluconobacter | cut -f1 | genprimers primer universe.fna output_prefix
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