[Cloning and expression of tryptophan synthetase gene (TRP5) in yeast]

TRP5, one of five genes required for tryptophan synthetase in S. cerevisiae, has been isolated on recombinant plasmids. A genomic DNA bank, containing the entire yeast genome was constructed by complete digestion of yeast 1412-4D DNA with restriction endonuclease BamH1, size fractionated by sucrose gradient (2-4 kb), and insertion of the fragments into the yeast shuttle vector pCN60. 9 recombinants plasmids capable of complementing trp5 mutations were isolated by transformation of yeast cell C9 (alpha, trp5, adel, ade6). The recombinant plasmids, containing 3.2 kb DNA fragments located TRP5 gene, were named pCN60 (TRP5). Tryptophan synthetase activity of transformants was 3-fold higher than that of original strain 1412-4D.