I'm designing species specific PCR probe (DNA hybridization probe) for a single gene in 10 different species as a diagnosis tool. I'm able to find DNA regions which are unique for the particular species, to design the probe. But in some species those region contain more than 75% GC, and their Tm lies around 70-80 degree Celsius (almost 15-20 degree Celsius greater than my primer).

I'm keeping the probe sequence length to a minimum of 17 bases to avoid high GC content in those species. But in that case I could find at least 75% similarity in DNA sequence with the probe (almost 12 bases), in other species as well as in other regions of the same species. Can we use this probe even though there is 75% similarity in sequence with other regions of DNA? If we can use, what should be done in these cases to run a PCR?

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