Summary
Yeast plasmids have been constructed that carry the Herpes simplex Virus type 1 (HSV-1) thymidine kinase (TK) gene which is functionally expressed in Saccharomyces cerevisiae. The expression of the TK gene appears to be due to transcriptional read-through from a yeast promoter that lies on the 3′ side of the HIS3 gene. The TK+ yeast possesses in vitro thymidine kinase activity which is absent in the original yeast strain. Yeast strains auxotrophic for thymidine monophosphate (dTMP) (tmp1) can grown on thymidine-containing medium after transformation with these plasmids. Tmp+, TK+ S. cerevisiae whose de novo synthesis of dTMP is inhibited with amethopterin plus sufanilamide is also capable of growth in thymidine. S. cerevisiae transformed with such plasmids is capable of incorporating thymidine and bromodeoxyuridine into DNA.
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References
Anderson EH (1946) Growth requirements of virus resistant Escherichia coli strain B. Proc Natl Acad Sci USA 32:120–122
Beggs JD, van den Berg J, van Ooyen A, Weissmann C (1980) Abnormal expression of chromosomal rabbit β-globin gene in Saccharomyces cerevisiae. Nature 283:835–840
Bolivar F, Rodriquez RL, Green PJ, Betlach MC, Heyneker HL, Boyer HW, Grosa JH, Falkow S (1977) The circular restriction map of pBR 322. In: Bukhari AI, Sharpiro JA, Adhya SL (eds) DNA insertion elements, plasmids and episomes. Cold Spring Harbor Laboratory, Cold Spring Harbor, New York, p 686–687
Clewell DB, Helinski DR (1970) Properties of a supercoiled deoxyribonucleic acid-protein relaxation complex and strand specificity of the relaxation event. Biochemistry 9:4428–4440
Cohen JD, Eccleshall TR, Needleman RB, Federoff H, Buchferer BA, Marmur J (1980) Functional expression in yeast of the Escherichia coli plasmid gene coding for chloramphenicol acetyltransferase. Proc Natl Acad Sci USA 77:1078–1082
Collins J, Hohn B (1978) Cosmids: A type of plasmid gene-cloning vector that is packageable in vitro in bacteriophage λ heads. Proc Natl Acad Sci USA 75:4242–4246
Enquist LW, van de Woude GF, Wagner M, Smiley JR, Summers WC (1980) Construction and characterization of a recombinant plasmid encoding the gene for the thymidine kinase of Herpes simplex type I virus. Gene 7:335–342
Fäth WW, Brendel M (1974) Specific DNA-labelling by exogenous thymidine-5′-monophosphate in Saccharomyces cerevisiae. Mol Gen Genet 131:57–67
Grivell AR, Jackson JF (1968) Thymidine kinase: Evidence for its absence from Neurospora crassa, and some other microorganisms, and the relevance of this to the specific labelling of deoxyribonucleic acid. J Gen Microbiol 54:307–317
Henikoff S, Tatchell K, Hall BH, Nasmyth KA (1981) Isolation of a gene from Drosophila by complementation in yeast. Nature 289:33–37
Hinnen A, Hicks JB, Fink GR (1978) Transformation of yeast. Proc Natl Acad Sci USA 75:1929–1933
Kiss GB, Cornish KV, Pearlman PE, Friesen JD (1980) The Herpes Simplex virus thymidine kinase gene is not expressed in Saccharomyces cerevisiae. Recombinant DNA Technical Bulletin 3:21–24
Klemperer HG, Haynes GR, Shedden WIH, Watson DH (1967) A virus-specific thymidine kinase in BHK 21 cells infected with Herpes Simplex Virus. Virol 31:120–128
Kunz BA, Barclay BJ, Game JC, Little JG, Haynes RH (1980) Induction of mitotic recombination in yeast by starvation for thymine nucleotides. Proc Natl Acad Sci USA 77:6059–6061
Laskowski W, Lehmann-Brauns E (1973) Mutants of Saccharomyces able to growth after inhibition of thymidine phosphate synthesis. Mol Gen Genet 125:275–277
Lennox ES (1955) Transduction of linked genetic characters of the host by bacteriophage PI. Virol 1:190–206
Little JG, Haynes RH (1973) DNA-specific labelling in yeast mutants. Genetics 74:S161
Little JG, Haynes RH (1979) Isolation and characterization of yeast mutants auxotrophic for 2′-deoxythymidine 5′-monophosphate. Mol Gen Genet 168:141–151
Mandel M, Higa A (1970) Calcium-dependent bacteriophage DNA infection. J Mol Biol 73:453–471
Maxam AM, Gilbert W (1977) A new method for sequencing DNA. Proc Natl Acad Sci USA 74:560–564
McKnight SL, Gavis ER (1980) Expression of the Herpes thymidine kinase gene in Xenopus laevis oocytes: An assay for the study of deletion mutants constructed in vitro.Nucl Acid Res 8:5931–5948
McKnight SL (1980) The nucleotide sequence and transcript map of the Herpes Simplex virus thymidine kinase gene. Nucl Acid Res 8:5949–5964
McNeil JB, Storms RK, Friesen JD (1980) High frequency recombination and the expression of genes cloned on chimeric yeast plasmids: Identification of a fragment of 2-μm circle essential for transformation. Current Genetics 2:17–25
Mercereau-Puijalon O, Lacroute F, Kourilsky P (1980) Synthesis of a chicken ovalbumin-like protein in the yeast Saccharomyces cerevisiae. Gene 11:163–167
O'Donovan GA (1978) Thymidine metabolism in bacteria (and how or how not to label DNA). In: Molineux I, Kohiyama M (eds) DNA synthesis: present and future. Plenum, New York, p 219–253
Panthier J-J, Fournier P, Heslot H, Rambach A (1980) Cloned β-galactosidase gene of Escherichia coli is expressed in the yeast Saccharomyces cerevisiae. Current Genetics 2:109–113
Southern EM (1975) Detection of specific sequences among DNA fragments separated by electrophoresis. J Mol Biol 98:503–517
Storms RK, McNeil JB, Khandekar PS, An G, Parker J, Friesen JD (1979) Chimeric plasmids for cloning of deoxyribonucleic acid sequences in Saccharomyces cerevisiae. J Bacteriol 140:73–82
Wagner MJ, Sharp JA, Summers WC (1981) Nucleotide sequence of the thymidine kinase gene of herpes simplex virus type 1. Proc Natl Acad Sci USA 78:1441–1445
Wickner RB (1974) Mutants of Saccharomyces cerevisiae that incorporate deoxythymidine-5′-monophosphate into deoxyribonucleic acid in vivo. J Bacteriol 117:252–260
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Communicated by G. Fink
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McNeil, J.B., Friesen, J.D. Expression of the Herpes simplex virus thymidine kinase gene in Saccharomyces cerevisiae . Mol Gen Genet 184, 386–393 (1981). https://doi.org/10.1007/BF00352510
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DOI: https://doi.org/10.1007/BF00352510