2 replies to this topic

[Su.J]

Hello,
I have been trying to amplify a 3'UTR region of an RNA virus. SO i initially made the cDNA and now doing end-point PCR to amplify the UTR region.My primers are to the ends of the UTR region along wiht the restriction sites attached to the prrimers.Total product size 616bp.
I have been getting a clear band in the b-actin control run in the same run, but not the band of my interest. but there are loads of primer dimers at the end!!any suggestions???is it just the annealing temperature?

[SabbathWarPig]

What annealing temperature are you using? Have you analysed your primer sequences for dimers and/or hairpin structures? and what is your melting temp of your primers? If not, a great program is oligo analyzer. just punch in your primer sequence and it gives you all the information you need

[SabbathWarPig]

a

Edited by SabbathWarPig, 30 August 2011 - 06:42 AM.