This proposal seeks to develop an immobilized cell reactor in which a recombinant DNA instability problem (plasmid instability) is overcome by selectively killing cells without recombinant DNA (without a plasmid). This is to be done using a toxin (colicin) to E. coli cells. A plasmid is to be used that confers resistance of these cells to colicin as well as a receptor site on the cell surface to this toxin. Colicin is to be attached to a column of sepharose beads and act as a binding agent for the E. coli. Those cells without the plasmid presumably will be killed and wash off the column while those cells with the plasmid will presumably stay attached to the column. The advantage of the above system to a suspension cell culture (fermentation) is twofold. - The toxin is not in solution and will not wash out of the reactor thus saving on the cost of the toxin. - The product of the bioreactor will not need to pass through a purification step to eliminate the toxin. The PI is considered to be well qualified to carry out the proposed research and I recommend funding of this proposal.
