The yeast Kluyveromyces lactis synthesizes a β-galactosidase (EC 220.127.116.11) which is inducible by lactose. We have isolated the gene that codes for this enzyme using recombinant DNA techniques. K. lactis DNA was partially digested with the restriction endonuclease Eco R1 and joined to Eco R1-digested pBR322 plasmid DNA using DNA ligase. A lac- mutant of Escherichia coil lacking the structural gene for β-galactosidase was transformed with ligated DNA. Three lac+ transformants containing recombinant plasmids were selected. Two of the plasmids (pK15 and pK17) contain four Eco R1-K. lactis DNA fragments having molecular weights of 2.2, 1.4, 0.55 and 0.5 × 106 daltons. The other plasmid (pK16) lacks the smallest fragment. E. coil carrying any of these plasmids produce β-galactosidase activity that has a sedimentation coefficient and immunological determinants that are nearly identical to K. lactis β-galactosidase and distinctly different from E. coli β-galactosidase. DNA-DNA hybridization studies show that the four Eco R1 fragments in pK15 hybridize to K. lactis but not to E. coil DNA.
|Number of pages||8|
|State||Published - Sep 1978|
Bibliographical noteFunding Information:
We are grateful to Dr . E . James and S . Moore for donating restriction enzymes and for technical advice, and to Drs . D . Pereira and R . Curtiss for sending us strain X2175 . We thank Drs . M . D . Barkley, L . R . Lacy, J . Lesnaw and R . L . Lester for their critical review of this manuscript . This research was supported by grants from the USPHS and Biomedical Research Support .
ASJC Scopus subject areas
- Biochemistry, Genetics and Molecular Biology (all)