DNA from uncultured organisms as a source of 2,5-diketo-D-gluconic acid reductases.

ABSTRACT: Total DNA of a population of uncultured organisms was extracted from soil samples, and by using PCR methods, the genes encoding two different 2,5-diketo-D-gluconic acid reductases (DKGRs) were recovered. Degenerate PCR primers based on published sequence information gave internal gene fragments homologous to known DKGRs. Nested primers specific for the internal fragments were combined with random primers to amplify flanking gene fragments from the environmental DNA, and two hypothetical full-length genes were predicted from the combined sequences. Based on these predictions, specific primers were used to amplify the two complete genes in single PCRs. These genes were cloned and expressed in Escherichia coli. The purified gene products catalyzed the reduction of 2,5-diketo-D-gluconic acid t

SUBMITTER: Eschenfeldt WH 

PROVIDER: S-EPMC93149 | biostudies-literature | 2001 Sep

REPOSITORIES: biostudies-literature