Vary, Patricia S.
M.S. (Master of Science)
Department of Biological Sciences
Bacterial genetics||Leucine||Bacillus megaterium||Amino acids--Synthesis
This thesis represents work done to determine the optimal conditions for transduction of Bacillus megaterium QM B1551 by bacteriophage MP13 and to use MP13 to initiate genetic studies of leucine biosynthesis in this bacterium. To determine the conditions which would increase the proportion of transducing particles, media, pH, temperature, aeration, multiplicity of infection and growth stage of the host were varied during propagation of the lysate. Conditions of the transduction procedure which were varied included: growth media of the recipient, phage inactivation time, selective media, pH of the selective media, incubation temperature and the use of antiserum. The results indicated a higher proportion of transducing particles were produced when the lysates were propagated in SNB broth, pH 8.0 at 30°C or 37°C with reduced aeration and a low multiplicity of infection. Recovery of transductants was increased when the recipient was grown in M broth at 30°C to late logarithmic stage, and the recipient was plated on an acidic selection medium with phage inactivated by 20 to 40 seconds of UV exposure. Increased recovery of transductants was observed when incubation was at 30°C but antiserum did not enhance recovery. After establishment of these optimal conditions for transduction, MP13 was used to analyse the leucine genes of 13. megaterium. Ten leucine mutations were found to be cotransducible and linked to ilv and phe markers. Biochemical and genetic analysis indicated the order of the genes to be ilv-leuA-leuC-leuB-phe.
Garbe, James C., "Development of a transducing system for Bacillus megaterium QM B1551 and the genetic analysis of leucine biosynthesis" (1981). Graduate Research Theses & Dissertations. 2180.
Northern Illinois University
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