So I want to choose the correct set of pair of primers to amplify the ORF of the gene that corresponds to amino acids in a protein. The start and stop codons are underlined. (I know that these need to be synthesised by the primer also for correct PCR technique)

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The boxed sets of nucleotides are the primer options I have at hand. I know that the DNA synthesis can only carry out from 5' to 3' and there is a forward and reverse primer. The forward primer should be easy enough but non of the options seem acceptable? Could someone please explain to me how to go about this problem?


closed as off-topic by MattDMo, AliceD, rg255, canadianer, March Ho Jul 2 '15 at 21:25

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  • $\begingroup$ So you've identified the ORF by marking the start and stop codons. You'll want a primer on either side of the ORF. You said none of the options seem ok, but why do you think that? $\endgroup$ – user137 Jun 27 '15 at 8:07

First of all, if this is a homework question, add that tag to it. And show what work you have done so far.

Secondly, primers for amplifications should lie on opposite strands. Primers are typed in 5'-to-3' direction (aka left-to-right on leading strand).

Appropriate primers will be: primer 4=GTG... and primer 5=GAA.... Note how those primers are always in 5'-3' orientation. That is how you order those.

Again, if this is homework (looks like it) it is very bad taste to just ask question without any work shown. It displays laziness and usually not respected behavior when asking for help.


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