Transfection of mouse cells with thymidine kinase gene of herpes simplex virus.
A mouse cell line (LP1-1) was established from the murine L cells deficient in thymidine kinase (L-M(TK-] by prolonged selective culture on the hypoxanthine-aminopterine-thymidine (HAT) medium following transfection with the thymidine kinase gene of herpes simplex virus type-I (HSVTK). Southern blot analysis has shown that the viral TK gene was integrated into one of the chromosomal loci by a single copy. From this established cell line, the 5-bromo-2-deoxyuridine (BrdU) resistant revertant was brought out at a frequency of 1 x 10(-6) and from these BrdU resistant revertants (LP1BU), one out of 1 x 10(5) cells could return to the HAT-resistant phenotype. The established LP1-1 cell line showed a typical biphasic nature of DNA synthesis as determined by the 3H-thymidine incorporation test. The activity of thymidine kinase was shown to be equivalent to that of the DNA polymerase-alpha when the whole nuclear fraction or the nuclear matrix were used for examination. These results indicate that the transfected viral TK gene can be expressed under the normal cell-cycle regulation and its gene product can act as a component of the multienzyme complex which is responsible for DNA replication.
Molecular Biology Unit, Memorial Sloan-Kettering Cancer Center, New York, NY 10021.
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CG
Lee
C G
CG
Kim
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Namkung
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SE
Lee
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Park
D000818:Animals; D002453:Cell Cycle / Q000502:physiology; D002460:Cell Line / Q000201:enzymology*; D004257:DNA Polymerase II / Q000378:metabolism; D015971:Gene Expression Regulation, Enzymologic; D007739:L Cells; D051379:Mice; D009097:Multienzyme Complexes / Q000378:metabolism; D018139:Simplexvirus / Q000201:enzymology* / Q000235:genetics; D013937:Thymidine Kinase / Q000235:genetics* / Q000378:metabolism; D014162:Transfection
D009097:Multienzyme Complexes; D013937:Thymidine Kinase; D004257:DNA Polymerase II