Discussion of all aspects of biological molecules, biochemical processes and laboratory procedures in the field.
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I have a relatively basic question regarding how to calculate my primer Tm although I have managed to confuse myself. I have designed two primers to amplify a target sequence of DNA and was required to engineer a restriction site to the 5' tails of each primer to allow for isolation of the amplified DNA for insertion to a plasmid. With regards to calculating the melting point of both primers, do I include the nucleotides on the 5' tails in my calculation or only use the nucleotides that match the target DNA.
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