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Calculation of Primer Melting Temp

Discussion of all aspects of biological molecules, biochemical processes and laboratory procedures in the field.

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Calculation of Primer Melting Temp

Postby sam2400 » Mon May 23, 2011 6:18 am

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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Postby JackBean » Mon May 23, 2011 8:09 am

You should use only those, which will anneal.

Cis or trans? That's what matters.
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