Use RT-PCR to verify after transient transfection a plasmid?transent transfection
Posted 13 September 2012 - 08:14 AM
Recently I transient transfected a plasmid which carry my target gene (size 7kb) in to mammalian cells (LO2) but I can't detect any protein changes through western blot.
So I try to use RTPCR to detect the mRNA level of my target gene if it rise up after transient.
But one of my labmates ask me how can I make sure the RTPCR has fold changes due to my target gene from mRNA but not the plasmid template in the cells.
Simply how can I make sure the RTPCR is amplifying mRNA but not DNA? (the target gene in the plasmid is translated region from cDNA)
Thanks in advance!
Posted 13 September 2012 - 09:01 AM
Edited by prabhubct, 13 September 2012 - 09:10 AM.
Posted 13 September 2012 - 09:02 AM
Posted 13 September 2012 - 09:13 AM
Posted 13 September 2012 - 09:18 AM
Posted 13 September 2012 - 07:24 PM