2002 Volume 35 Issue 5 Pages 474-478
There are two kinds of mating types in yeast cells, a-type and α-type, and they can make a diploid, which carries both the genes originated from a-type and α-type. We propose a new method to obtain a larger size of combinatorial libraries by mating these yeast cells. By transforming libraries of genes of heavy and light chains to a- and α-types respectively, for example, it should be possible to obtain a larger size of libraries of antibodies by mating them. To confirm our method, the expression vectors containing heavy chain gene (μ) and light chain genes (κ and λ) of human antibodies were transformed into a-type and α-type strains of S. cerevisiae, respectively, and these transformants were mated by mixing the cultured cells of both types. The effects of media, cell concentration and the ratio of a-type cells to α-type cells on the mating efficiency were studied. The direct contact between a- and α-type cells was essential to initiate mating of cells, and the probability of contact among cells determined the mating efficiency, when cells are well nourished. Stable diploids were formed by the combination of plasmids of the 2 μm types, as well as the combination between the 2 μm type and the ARS1 type.
