Hello all,
I am trying to transfect my bacmid to sf-21 cell line for baculovirus generation and I am facing a bacterial contamination in the transfected well. I used alkaline lysis method to isolate my bacmid. Precipitated it with isopropanol and dissolved in tris-EDTA(0.1mM). What could be the possible reason and how should I get rid of it. After transfection the cells are incubated with SFM+1X antibiotic media.
Michael J. Benedik
I find it hard to imagine that any host E. coli cells survived alkaline lysis and isopropanol precipitation. I wonder whether the bacterial contamination is coming from elsewhere. But if you want to be sure, you can always take the plasmid DNA and incubate 10 minutes or so at 65C or even 80C. E. coli does not survive above 50C.
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