designing primers( selecting target sequence/amplicon design) - (Jan/06/2014 )
Okay this sounds a bit dumb but all these are new to me, so please help in anyway you can.
I'm trying to design a new primers (let's say for a bacteria). I know for amplicons you're suppose to select 75-100bp, avoid secondary structures etc...
but how do you go about selecting the target sequence? Is there some kind of database where you can see the dna sequence of a bacteria and can select a target sequence/amplicons on that? and thus the primers.
I'm sorry if I'm not being clear...please any help would be much appreciated.
This sounds like homework to me....... so my answer won't be complete, but should be enough to get you started.
Have a look for genbank.
What type of PCR are you doing (for what purpose)? qPCR (real time) or standard?
It's not homework I assure you. I just never design primers before and only study in a classroom (and they only tell you how to design primers , not exactly how to do it in real life ) instead of practical experience. and practical experience with PCR only and with already prepared primers.
And It's for qPCR.
There are databases of validated (published) qPCR primers out there, but I don't know the websites off hand. Have a look at the Nucleic Acids Research database issue.
Check out PrimerBlast on the NCBI website.