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Selection of spontaneous mutants ofYarrowia lipolytica by inositol-less death

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Journal of Industrial Microbiology

Summary

A procedure was developed for the selection of spontaneous mutants of the yeastYarrowia lipolytica. An inositol-requiring mutant of a wild-typeY. lipolytica, YB 3-122, was derived by mutagenesis and screening. The mutant had a reversion frequency of less than 6×10−9. A mutant selection procedure based on inositolless death was then developed using this mutant strain. The selection procedure killed growingY. lipolytica cells and enriched for mutants yielding cultures that consisted of 60–98% spontaneous mutants after two rounds of inositol-less death. The procedure enriched for four classes of mutants, strains that were auxotrophic, metabolite analog sensitive, temperature sensitive, or unable to grow on citric acid as the sole carbon source. Since strain YB 3-122 is now available to yeast researchers, inositol-less death will be useful for the routine isolation of spontaneous mutants ofY. lipolytica.

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References

  1. Ahearn, D.G., S.P. Meyers and R.A. Nichols. 1968. Extracellular proteinases of yeast and yeastlike fungi. Appl. Microbiol. 16: 1370–1374.

    PubMed  Google Scholar 

  2. Akiyama, S., T. Suzuki, Y. Sumino, Y. Nakao and H. Fukuda. 1972. Production of citric acid fromn-paraffins by fluoroacetate-sensitive mutant strains ofCandida lipolytica. Agric. Biol. Chem. 36: 339–341.

    Google Scholar 

  3. Barth, G. and H. Weber. 1983. Genetic studies on the yeastSaccharomycopsis lipolytica. Z. Allg. Mikrobiol. 23: 147–157.

    PubMed  Google Scholar 

  4. Barth, G. and H. Weber. 1984. Use of nystatin for random spore selection in the yeastSaccharomycopsis lipolytica. Z. Allg. Mikrobiol. 24: 125–127.

    PubMed  Google Scholar 

  5. Beckerich, J.M., H. Blanc, M. Lambert and L. Louvel. 1980. Studies on glycerol and inositol auxotrophic mutants inSaccharomycopsis lipolytica. Adv. Biotechnol. 4: 471–477.

    Google Scholar 

  6. Bigelis, R. 1985. Primary metabolism and industrial fermentations. In: Gene Manipulations in Fungi (J.W. Bennett and L.L. Lasure, eds.), pp. 357–401, Academic Press, New York.

    Google Scholar 

  7. Bigelis, R. and K.A. Black. 1985. Selection ofYarrowia liplytica mutants by inositol-less death. Abstr. Annu. Mtg. Am. Soc. Microbiol. K119.

  8. Culbertson, M.R. and S.A. Henry. 1975. Inositol-requiring mutants ofSaccharomyces cerevisiae. Genetics 80: 23–40.

    PubMed  Google Scholar 

  9. Esser, K. and U. Stahl. 1976. Cytological and genetic studies of the life cycle ofSaccharomycopsis lipolytica. Mol. Gen. Genet. 146: 101–106.

    PubMed  Google Scholar 

  10. Fink, G.R. 1970. The biochemical genetics of yeast. Methods Enzymol. 17A: 59–78.

    Google Scholar 

  11. Fukuda, H., T. Suzuki, S. Akiyama and Y. Sumino. 1974. Microbiological production of citric acid using a paraffin substrate. U.S. Patent No. 3,799,840.

  12. Henry, S.A., T.F. Donohue and M.R. Culbertson. 1975. Selection of spontaneous mutants by inositol starvation in yeast. Mol. Gen. Genet. 143: 5–11.

    PubMed  Google Scholar 

  13. Henry S.A., K.D. Atkinson, A.I. Kolat and M.R. Culbertson. 1977. Growth and metabolism of inositol-starvedSaccharomyces cerevisiae. J. Bacteriol. 130: 472–484.

    PubMed  Google Scholar 

  14. Holliday, R. 1962. Selection of auxotrophs by inositol starvation inUstilogo maydis. Microbial Gen. Bull. 13: 28–30.

    Google Scholar 

  15. Lester, H.E. and S.R. Gross. 1959. Efficient method of selection of auxotrophic mutants ofNeurospora. Science 139: 572.

    Google Scholar 

  16. Mattey, M. and M. Morgan. 1978. Secretion of extracellular lipases byCandida lipolytica. Biochem. Soc. Trans. 6: 426–428.

    PubMed  Google Scholar 

  17. Moat, A.G., N. Peters, Jr. and A.M. Srb. 1959. Selection and isolation of auxotrophic yeast mutants with the aid of antibiotics. J. Bacteriol. 77: 673–677.

    PubMed  Google Scholar 

  18. Nga, B.H., H. Heslot, C.M. Gaillardin, P. Fournier, K. Chan, Y.N. Chan, E.W. Lim and P.C. Nai. 1988. Use of nystatin for selection of tributyrin non-utilizing mutants ofYarrowia lipolytica. J. Biotechnol. 7: 83–86.

    Google Scholar 

  19. Ogrydziak, D.M. and S.J. Scharf. 1982. Alkaline extracellular protease produced bySaccharomycopsis lipolytica CX161-1B. J. Gen. Microbiol. 128: 1225–1234.

    PubMed  Google Scholar 

  20. Orgrydziak, D.M., A. L. Demain and S.R. Tannenbaum. 1977. Regulation of extracellular protease production inCandida lipolytica. Biochim. Biophys. Acta 497: 525–538.

    PubMed  Google Scholar 

  21. Ogrydziak, D., J. Bassel, R. Contopoulou and R. Mortimer. 1978. Development of genetic techniques and the genetic map of the yeastSaccharomycopsis lipolytica. Mol. Gen. Genet. 163: 229–239.

    Google Scholar 

  22. Takeda Chemical Industries, Ltd. 1970. Procede de production d'acide citrique. Belgian Patent No. 744416.

  23. Tobe, S., T. Takami, S. Ikeda and K. Mitsugi. 1976. Production and some enzymatic properties of alkaline proteinase fromCandida lipolytica. Agric. Biol. Chem. 40: 1087–1092.

    Google Scholar 

  24. Pfizer and Co., Inc. 1970. Citric acid prepared by growing candida yeasts on alkanes or alkenes. British Patent No. 1,203,006.

  25. Pfizer, Ltd. 1974. Citric acid production. British Patent No. 1,369,295.

  26. Snow, R. 1966. An enrichment method for auxotrophic yeast mutants using the antibiotic ‘nystatin’. Nature 211: 206–207.

    PubMed  Google Scholar 

  27. Ulaszewski, S., J.R. Woodward and V.P. Cirillo. 1978. Membrane damage associated with inositol-less death inSaccharomyces cerevisiae. J. Bacteriol. 136: 49–54.

    PubMed  Google Scholar 

  28. Yano, K., M. Kawamura and M. Takagi. 1981. Enrichment ofn-alkane assimilation-deficient mutants ofCandida yeast by synergistic effect of nystatin and pyrrolnitrin. Agric. Biol. Chem. 45: 1017–1018.

    Google Scholar 

  29. Yarrow, D. 1972. Four new combinations in yeasts. Antonie van Leeuwenhoek 38: 357–360.

    PubMed  Google Scholar 

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Bigelis, R., Black, K.A. Selection of spontaneous mutants ofYarrowia lipolytica by inositol-less death. Journal of Industrial Microbiology 4, 435–440 (1989). https://doi.org/10.1007/BF01569639

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