expression of short RNAs - (May/08/2007 )
HI everyone,
I'm trying to express short single stranded RNAs (~15-40 nts) in mammalian cell cultures. I started out with just a random sequence I made up. It seems that the expression of my single stranded RNA from a plasmid affects the cells negatively. (it was unable to express high levels of a EGFP reporter gene that I cotransfected.) I am not sure why expression of theses short RNAs would affect the cell. The ssRNA has some complementary to some genomic DNA and blasting the the sequence does not show any complementary to any human mRNA. Any one have any idea whats going on? Also if anyone know or has advice about a sequence I can express in the cell that would be very helpful.
Thanks
Why would you like to express a scramble ssRNA in your cells? For control purposes?
Also, you mentionned a mammalian cell line. Is it a human cell line? Because if not, your ssRNA could have some complementarity with the mRNAs of the cell line's specie.
Also, you mentionned a mammalian cell line. Is it a human cell line? Because if not, your ssRNA could have some complementarity with the mRNAs of the cell line's specie.
I'm expressing the ssRNA in human cells (hela and 293). The ssRNA is meant as a control. I have one sequence that is suppose to form a duplex with another sequence in the cell and another one is not suppose to form a duplex (the control) but I have been unable to construct a control sequence that does not affect the transfection effenency of the cell.