smears instead of bands
Posted 23 June 2004 - 02:27 AM
i have successfully run pcr reactions looking for ucp-2 mRNA for 4 years, however, in the last months the pcr has stopped working. instead of easily repeatable bands, i am getting no bands and just a smear down the whole lane.
i use abgene's pcr reddy mix mastermix tubes which have always been fine, and reverse transcribe using 10 units of RT which again has always been fine
i get some more success with my actin bands, this led me to believe it was my ucp-2 primers which were the problem [as positive control was not working either] but some actin bands also disappear, and now my positive control has started to give bands again..... HELP!!!!
Posted 05 August 2004 - 09:04 PM
Edited by SVTX, 05 August 2004 - 09:04 PM.
Posted 06 August 2004 - 09:42 AM
i tried changing primers(to qiagen .inc) and renewed all reagents for PCR,got some success
also take care all this are RNase free b4 u start working