CLONING, CONSTRUCTION OF PROKARYOTIC EXPRESSION VECTOR AND EXPRESSIONOF ESCHERICHIA-COLI CYTOSINE DEAMINASE GENE

Cytosine deaminase gene of Escherichia coli strain H-30 was cloned, an d its initiation codon of 'GTG' was mutated to 'ATG' by PCR. Prokaryot ic recombinant expression vector pBV220-CD was constructed. Clone with high enzyme activity were selected by detecting their specific activi ty of cytosine deaminase. 5-FC(5-FC, 5-fluorocytosine) could induce th e lethal toxicity to cells containing active CD gene. DNA sequence ana lysis indicated that there were 16 altered bases and 5 of them resulte d in the alteration of amino acids in predicted peptide by comparing D NA sequence of the clone H-30-CD-11 with high enzyme activity with CD gene reported in Gene Bank.