I have 10 closely related sequence, and I want to create a common primer for that, how can I create common primer ?

Question

[bioinformaticssrm2011]

Is there any software by which user can enter all the closely related sequence in it, and software create the common primer?

[bioinformaticssrm2011]

I have done MSA using mega6 software, than how can I select primer pairs that span gaps of low similarity?

[Michael]

Can't you use Jalview to visualize the alignment and detect identical and variable regions?

[bioinformaticssrm2011]

I have used the Jalview to visualize it, but there is not much identical region are available.

[Michael]

Possibly you can do it using two primer pairs, will only require 2 reactions. Look for groups of sequences that are identical within the whole set, a group of 5 for example. If you do not have any identical stretches, then it will render the task of finding universal primers impossible, and you would have to make 10 pairs and 10 reactions.

Answer 1

[Michael]

Do a multiple sequence alignment and find regions of high conservation between all sequences, then try to find primer pairs for the consensus sequence, try to select primer pairs that span gaps of low similarity. Your intention is most likely to amplify and then determine the species which are present, therefore you need a variable region which is located between conserved sequences. If you are looking for bacterial 16s rRNA sequences, this has already been solved and there are universal primers for this purpose.