how to get rid of the DNA within RNA extraction - (Sep/05/2007 )
Hi guys:
I am running real time RT-PCR with RNA extracted from tissue sample. Although on column DNase treatment is included in the RNA extraction protocol.( I used Ambion RecoverAll kit) . Positive band still showed up for no-RT control. My feeling is the crude RNA needs secondary DNase treatment, any good suggestion on that?
Thanks!
-whack-
Do you check the PCR without template (use H2O instead of RT products)? If positive band still showed up without template, the comtemination of pipptors, solutions should be considered.
A cracked air bubble of the PCR product may caused the pipptors conteminated.
-zhongmindai-
QUOTE (zhongmindai @ Sep 5 2007, 08:14 PM)
Do you check the PCR without template (use H2O instead of RT products)? If positive band still showed up without template, the comtemination of pipptors, solutions should be considered.
A cracked air bubble of the PCR product may caused the pipptors conteminated.
A cracked air bubble of the PCR product may caused the pipptors conteminated.
yes, I did run a non template control. it is negative. And also my non amplification control is negative(without taq). Only this no-RT control and my sample showed positive results.
-whack-
turbo DNase from ambion gave better results. and as it's bead-atache, you can easily get rid of the protein.
-fred_33-