How to design or make a primer for PCR?

Question

I need to design a primer to test for a large group of viruses and just wanted to know how to make a primer, what things i need inorder for it to work

Ok thanks for the input so far it was great.
But how do you make the solution of the primer, primers for PCR are in liquid form, how do you go from a sequence to that liquid form

Answer 1

Primers must be complementary to the sequence you hope to ampify. They must also be in pairs so that you amplify the region of DNA between the pairs.

You want minimal secondary structure in an individual primer (unlikely to form a hairpin), you don't want it to be comlementary with the second primer in the pair or you get "primer dimers" instead of having them anneal to your DNA of interest.

Try to avoid GC content that is too high so that your melting temperature is reasonable. Length is often 20 bases or thereabouts, but can vary considerably.

And if you really want to know how to make one, look up a primer program on the web, input your sequence, and have it suggest reasonable pairs for you to try. The Whitehead Institute at MIT has one (I think you need to register there), but there are lots of other that anyone can use.

Answer 2

You need a sequence that will be present in the viruses, 11 to 15 bp long. You have to consider if these are ssDNA viruses or RNA viruses. Ideally, the sequence should be unique to the virus group.

Answer 3

Hi here I made some videos to answer for your question:

https://www.youtube.com/watch?v=l6aiunDTOJs


https://www.youtube.com/watch?v=rbvvvLWE8jI


https://www.youtube.com/watch?v=zZqjH1IibCs


https://www.youtube.com/watch?v=0vdbf4F90_o


https://www.youtube.com/watch?v=dnmZFOADwI8


https://www.youtube.com/watch?v=3I3l5xxZscg


https://www.youtube.com/watch?v=LGb6y4xn93w