i've been trying to use pmiR-report vector to clone a 2.7kb i3'utr and my problem is that even if I'm getting cells with the clone, all of them have any problem. The first time I got the colonies I find out that in one i had lost the luciferase gene, the second time it was the CMV to be lost and two days ago i found a plasmid which had like another extra 3kb in their sequence.
I've worked with other vectors and it seems to me that the error ratio here is much more higher that as usual!
Does anyone have an idea about what's going on? i thought that maybe it would be a problem with the compatibility of the e.coli strains i was using, maybe something about recombination. I've used JM109 from promega the first time, and now i was using DH10B. I haven't found any restrictions about the e.coli strains in the manual so right now i'm pretty desperate.
Edited by losybelle, 30 January 2012 - 11:46 PM.