I have a question regarding how do we know what temperature should we choose for the gradient in the pcr machine. For example, I have primer with average temperature= 57.4. I need to use 6 different degree of temperature. So how do i know what temperature should i choose?
Rajitha Wickramasinghe
In PCR, the annealing temperature is crucial for primer binding. Typically, it's chosen a few degrees below the primer's melting temperature (Tm). You can try a range around 57.4°C, perhaps 2-5°C lower and higher, to find the optimal temperature for efficient primer binding during annealing. Running a gradient PCR with a temperature range can help you identify the most suitable annealing temperature for your specific primers.
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