Designing diagnostic primers

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Tool 4 output description

Subject to positive identification, the ten longest unique portions of an isolate’s genome are deposited in the output file (*.candidate_unique.fa).

While these stretch are unique when compared with the reference library and the other subject isolates, it may still be possible that these stretches do appear in other organisms that were not included in the analysis. It is important to bear this in mind when designing diagnostic primers.

It is also worth considering the context of the unique portion when selecting which portion use for primer design. Will the site of the primers be stable?

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NCBI Primer BLAST

To design diagnostic primers NCBI’s Primer-BLAST tool is suggested. This will also check primer pair specificity.

Specify the ‘nr’ database. You can also specify the organism(s) that you wish to check for specificity, for example, ‘bacteria (taxid:2)’, or ‘Lactobacillales (taxid:186826)’ or ‘Lactobacillus (taxid:1578)’.