Archive for June, 2010

DNA Trasfection of Cells

Our vision at Aldevron is to be a world-class global contract services organization. To achieve this, we need to understand what our clients need and deliver what they want. One of our services is expression screening in insect cells. We offer a range of options to suite the clients needs. A typical screening project includes multiple variants, such as amino acid changes or truncations, of a single gene. We are able to screen multiple variants and culture conditions in parallel. There are a couple of options for expression screening in insect cells. The first option is to screen for expression via baculovirus infection. We can either start with a recombinant transfer vector supplied by the client or we can clone the gene of interest into a baculovirus transfer vector. Next, we generate a high-titer baculovirus stock and determine the viral titer. We then infect 10ml Sf21 cultures for recombinant protein expression. The second option is to screen for expression via direct transfection. This allows us to go from plasmid to protein in as little as 48 hours. We start by cloning the gene of interest into an expression vector designed to enable robust expression in insect cells without the need to produce recombinant baculovirus. Next we transfect 10ml Sf9 cultures with the recombinant plasmids. 48-72 hours post-transfection, we harvest the cells. Because generating a recombinant baculovirus can require anywhere between 2-4 weeks of time, the direct transfection method offers a …