Analysis of a method for selecting temperature-sensitive mutants of BHK cells

Abstract

A procedure for the isolation of temperature sensitive mutants of BHK cells is described. Mutagenized cells were synchronized in G1 by serum starvation at 33 degrees, and, following release, shifted to 37.5 degrees in the presence of FUdR, to kill cells entering the DNA synthetic phase. Eight independently mutagenized series of cells were carried through four cycles of this selection, and surviving cells were tested for ability to grow at 33 degrees and 39.5 degrees after each cycle. The results suggest that such procedure is effective in enriching for non-leaky ts mutants and favors the isolation of mutants which at 39.5 degrees are inhibited in the processes necessary for DNA synthesis and lose viability rapidly. The effectiveness of repeated selection cycles and the general characteristics of the ts mutants isolated by this method were also evaluated.