A member of our lab is using Primer-Blast to find what DNA her pairs of primers amplify in the NT nucleotide database. She designed degenerated primers and the number of possible pairs goes up to 256. She does not want to do that many primer-blasts by hand, obviously, and Primer-Blast apparently cannot use degenerate primers. I am therefore looking for a tool to automate these primer blasts.
Is there a command-line interface tool that would, given a local database of nucleotide sequences, return which sequences would amplify with a given pair of primers? It does not look like Primer-Blast can be downloaded for local and I don't think Primer3 can be used for that purpose.
My idea is to automatically generate all possible degenerate primer pairs and then run the CLI tool on them. If a program was able to directly use degenerate primers, that would be even awesomer :)
Thanks for your suggestions!
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Eric Normandeau
primerTree is the closest thing I've found to a command-line interface for Primer-BLAST. Here is the R package on CRAN. The primer_search command can use primers with IUPAC codes.
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Cameron Turner •
ThermonucleotideBLAST will search degenerate PCR primers against a fasta file (or BLAST-formatted nucleotide sequence database).
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JasonGans •
gfServer pcrDirect fPrimer rPrimer file(s).nibthere is also a standalone version of ncbi epcr: http://www.ncbi.nlm.nih.gov/tools/epcr/
Pierre, there are quite a bunch of tools listed at your first URL. None contain 'pcr' or 'primer' and the two that contain 'blast' are not what I am looking for. Would you have specific names for the tools you are thinking about?
see http://hgdownload.cse.ucsc.edu/admin/exe/linux.x86_64/FOOTER may be you'll have to compile it from the sources ?
Hi I've found the EMBOSS tool "primersearch" to be the most straightforward - for whatever reason they don't call it an in-silico pcr tool but that's what it does.
Thank you, I'll give primersearch a try.
Silica searches (multiple) PCR primers across entire genomes using primer3's thermodynamic calculations. It's an installation-free, open-source in-silico PCR web application: https://www.gear-genomics.com/silica/
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There is a tool called primertarget that does exactly what you need. http://www.genewide.com/
Eric, did primersearch solve your problem? I have the same question as you and haven't been able to find a good solution.