PMID:6315690
| Citation |
Iwakura, M, Shimura, Y, Sakai, T and Tsuda, K (1983) Purification of dihydrofolate reductase amplified in Escherichia coli K-12. J. Biochem. 94:1021-4 |
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| Abstract |
The Escherichia coli strain carrying pTP 6-10 which was constructed in our previous work (Iwakura, M., et al. (1983) J. Biochem. 93, 927-930) produces more than 400-fold dihydrofolate reductase as compared with the strain without the plasmid. Dihydrofolate reductase was highly purified from the cell-free extract of the plasmid strain simply by two steps; ammonium sulfate fractionation and ion-exchange chromatography. By 10-fold purification, the enzyme was essentially homogeneous as judged by polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulfate. The restriction map of pTP 6-10 was also determined and the plasmid was shown to have an Ava I, an EcoR I, a Pst I, a Pvu I, and a Pvu II site. Our results indicate that the plasmid strain is suitable as a source of the enzyme and that plasmid pTP 6-10 is promising as a versatile plasmid vector for efficiently yielding the product of the cloned gene. |
| Links | |
| Keywords |
Base Composition; Base Sequence; DNA Restriction Enzymes; Escherichia coli/enzymology; Escherichia coli/genetics; Gene Amplification; Mutation; Plasmids; Tetrahydrofolate Dehydrogenase/genetics; Tetrahydrofolate Dehydrogenase/isolation & purification |
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