Abstract
Antimetabolite-resistant and auxotrophic mutants of the hyperthermophilic bacterium Thermotoga neapolitana were isolated to provide strains with genetic backgrounds amenable to genetic analyses. Norleucine, azaleucine, 4-nitropyridine-N-oxide, and 3-amino-1, 2, 4-triazole did not affect growth, while 5-fluorouracil (5 μg/ml), 5-methyltryptophan (250μg/ml), 6-azauracil (100 μg/ml), and 4-fluorophenylalanine (30 μg/ml) inhibited growth at the indicated minimum inhibitory concentrations. The effect of 5-fluorouracil was analyzed and found to be bacteriostatic. These inhibitors were used to select spontaneously arising resistant mutants. In addition, auxotrophic mutants requiring leucine, tryptophan, adenine, and histidine were isolated following mutagenesis with ethyl methanesulfonate. Six other auxotrophs with undefined growth requirements were also isolated. These strains will be useful for the development of genetic methods for T. neapolitana.
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References
Achenbach-Richter L, Gupta R, Stetter KO, Woese CR (1987) Were the original eubacteria thermophiles? System Appl Microbiol 9:34–39
Adelberg EA (1958) Selection of bacterial mutants which excrete antagonists of antimetabolites. J Bacteriol 76:326
Belkin S, Wirsen CO, Jannasch HW (1986) A new sulfur-reducing, extremely thermophilic eubacterium from a submarine thermal vent. Appl Environ Microbiol 51:1180–1185
Bouthier de la Tour C, Portemer C, Huber R, Forterre P, Duguet M (1991) Reverse gyrase in thermophilic eubacteria. J Bacteriol 173:3921–3923
Childers SE, Vargas M, Noll KM (1992) Improved methods for cultivation of the extremely thermophilic bacterium Thermotoga neapolitana. Appl Environ Microbiol 58:3949–3953
Harriott OT, Huber R, Stetter KO, Betts PW, Noll KM (1994) A cryptic miniplasmid from the hyperthermophilic bacterium Thermotoga strain RQ7. J Bacteriol 176:2759–2762
Huber R, Stetter KO (1992) The order Thermotogales. In: Balows A, Trüper HG, Dworkin M, Harder W, Schleifer KH (eds) The prokaryotes, 2nd edn. Springer, Berlin Heidelberg New York, pp 3809–3815
Huber R, Langworthy TA, König H, Thomm M, Woese CR, Sleytr UB, Stetter extremely thermophilic eubacteria growing up to 90C. Arch Microbiol 144:324–333
Jannasch HW, Huber R, Belkin S, Stetter KO (1988) Thermotoga neapolitana sp. nov. of the extremely thermophilic eubacterial genus Thermotoga. Arch Microbiol 150:103–104
Londei P, Altamura S, Huber R, Stetter KO, Cammarano P (1988) Ribosomes of the extremely thermophilic eubacterium Thermotoga maritima are uniquely insensitive to the miscoding-inducing action of aminoglycoside antibiotics. J Bacteriol 170: 4353–4360
Matsumura M, Aiba S (1985) Screening for thermostable mutant of kanamycin nucleotidyltransferase by the use of a transformation system for a thermophile, Bacillus stearothermophilus. J Biol Chem 260:15298–15303
Miller JH (1972) Experiments in molecular genetics. Cold Spring Harbor Laboratory, Cold Spring Harbor, New York
Wachsman JT, Hogg L (1964) Use of 5-fluorouracil for the isolation of auxotrophic mutants of Bacillus megaterium. J Bacteriol 87:1137–1139
Wiegel J (1986) Methods for isolation and study of thermophiles. In: Brock TD (ed) Thermophiles: general, molecular, and applied microbiology. Wiley-Intersience, New York, pp 17–37
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Vargas, M., Noll, K.M. Isolation of auxotrophic and antimetabolite-resistant mutants of the hyperthermophilic bacterium Thermotoga neapolitana . Arch. Microbiol. 162, 357–361 (1994). https://doi.org/10.1007/BF00263784
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DOI: https://doi.org/10.1007/BF00263784