[Cloning, sequence analysis of deacetoxycephalosporin C synthetase/hydroxylase gene cefEF]

Chromosomal DNA preparations were made form the fungi C. acremonium and A. chrysogenum respectively by a method using 1-Chloro methyl benzene solution. Then the cefEF gene of deacetoxycephalosporin C synthetase/hydroxylase was amplified by polymerase chain reaction. The cefEF gene could only be amplified from the chromosomal DNA of A. chrysogenum. This fragment about 1.0 kb was cloned into pGME-T vector and then sequenced. The sequence indicates that the cloned cefEF gene contains 999 nucleotides encoding for 333 amino acids and there are three bases and two amino acids different from those that have been reported. Both of the two amino acids are not in active site. The result also indicates that the reported C. acremonium or A. chrysogenum generating deacetoxycephalosporin C synthetase/hydroxylase corresponds with A. chrysogenum but not C. acremonium in China.