This program designs PCR primers for a multiplex of given target regions of a DNA sequence in a FASTA file.
The script takes several arguments:
| Parameter | Description |
|---|---|
| region_file (required) | The path to the primer design region file. This file should have three columns: name, start position, and end position (1-based). It can be in either tsv or xlxs format. |
| input_file (required) | The path to the input FASTA file. |
| target_tm | The desired melting temperature (Tm) for the primers. Default is 60. |
| primer_len | The desired length of the primers. Default is 20. |
| product_size_min | The desired min size for the PCR product. Default is 400. |
| product_size_max | The desired max size for the PCR product. Default is 800. |
| ret | The maximum number of primer pairs to return. Default is 100. |
| Q5 | A boolean indicating whether to use NEB Q5 hotstart polymerase settings for primer3. Default is True. |
| background | The path to the mispriming library FASTA file. Default is an empty string. |
| output | The name of the output file. Default is 'MultiPlexPrimerSet'. |
| eval | The maximum number of primer sets to evaluate. Default is 10000. |
The main function of the script is 'design_primers', which takes the input FASTA file, start and end positions of a target region, and the arguments specified using argparse, and returns the best primer set for that region as a list of dictionaries containing information about the primer pairs.
The 'design_primers' function performs the following steps:
- Parses the input FASTA file and extracts the target region sequence.
- Sets up the primer3 input parameters based on the specified arguments.
- Runs primer3 to design the primers.
- Extracts information about the primer pairs.
- Determines the best set of multiplex primers.
Installation
pip install getMultiPrimerSet
Example usage
getMultiPrimerSet --region_file "./example/primer_regions.tsv" --input_file "./example/ref.fasta" --target_tm 65 --primer_len 20 --product_size_min 400 --product_size_max 800 --ret 100 --eval 1000 --Q5 --background "" --output "example"
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Metadata
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Release files for getMultiPrimerSet 1.0.2
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