I want to determine one contaminant exsit or not in my cultivation by PCR. However, I am not sure is the primers targeted in 16S sequence designed by NCBI specific enough to support my conclusion.
And, is that possible to choose some other specific gene according to RNAseq results?
Assuming you can grow pure cultures of your contaminant versus species of interest, perform a genomic DNA prep and test each set of primers on the various preps using PCR. Check for cross reactivity between primer sets. You can also play around with the annealing temp in order to make your primers more specific.
How about differences in restriction sites? Can you identify restriction sites present in you contaminant that are not present in your cultivar (or visa-versa)? Fairly inexpensive way to extend a PCR analysis.
You can taget specific gene or genetic element like IS, transposon
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