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Adaptation-Mar_24-31

Adaptation-Mar_24-31 - Environmental Pollution Microbiology...

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Environmental & Pollution Microbiology Spring 2010 Environmental Adaptation [Brock 11, chapter 10; Brock 12, chapter 11] (1.) QUESTION FOR DISCUSSION A. Toluene as a model contaminant. What happens when it is introduced into an aquifer? B. Concept of adaptation . Adaptation can involve population changes and individual regulon activation or derepression. The ultimate basis for individual and population changes are genetic changes .
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(2.) Dennis Focht’s experiments to obtain a chlorobenzene degrader A. references : Krockel, L., and D. D. Focht. 1987. Construction of chlorobenzene-utilizing recombinants by progenitive manifestation of a rare event. Appl. Environ. Microbiol. 53:2470-2475; Carney, B. F., L. Krockel, J. V. Leary, and D. D. Focht. 1989. Identification of Pseudomonas alcaligenes chromosomal DNA in the plasmid DNA of the chlorobenzene-degrading recombinant Pseudomonas putida strain CB1-9. Appl. Environ. Microbiol. 55:1037-1039. B. 2 starting strains P. alcaligenes C-O grows on 3-chlorbenzoate (15 kb plasmid); P. putida R5-3 grows on toluene (55 kb plasmid)
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C. Experimental set-up Chemostat 1 with strain C-O fed benzoate; cheomstat 2 with R5-3 fed toluene; chemostat 3 provided with chlorobenzene D. Outcome Turbidity detected in chemostat 3; medium turned bright yellow; chloride was released; obtained strain CB1-9 with phenotypic chartacteristics of P. putida ; CB1-9 carried 33 kb plasmid which hybridizes strongly to 55kb plasmid of R5-3, as well as having a band that hybridizes to chromosomal DNA of C-O. E. How did this new strain come about? What genetic events were involved? [Fig. 1; Refer to Hwk Problem]
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C. Experimental set-up Chemostat 1 with strain C-O fed benzoate; cheomstat 2 with R5-3 fed toluene; chemostat 3 provided with chlorobenzene D. Outcome Turbidity detected in chemostat 3; medium turned bright yellow; chloride was released; obtained strain CB1-9 with phenotypic chartacteristics of P. putida ; CB1-9 carried 33 kb plasmid which hybridizes strongly to 55kb plasmid of R5-3, as well as having a band that hybridizes to chromosomal DNA of C-O. E. How did this new strain come about? What genetic events were involved? [Fig. 1; Refer to Hwk Problem]
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You have just finished a long series of experiments with Dennis Focht’s strains R5-3 (the toluene degrader), C-0 (the chlorobenzoate degrader) and CB1-9 (the putative recombinant that is capable of chlorobenzene degradation). From your experimental work you have concluded that CB1-9 could not have arisen from either transduction (you could find no evidence for a transducing phage) or transformation (neither R5-3 nor C-0 become competent under any set of growth conditions that you tried). Homework Problem
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Your most recent set of experiments consisted of the following: 1.) You “genetically tagged” the resident plasmid of C-0 with a tetracycline (Tc) resistance gene and found through a series of conjugation experiments with several different recipient strains that the resident plasmid of C-0 is a conjugal plasmid since resistance to this antibiotic was acquired by the recipients following the conjugation experiments; 2.) You “genetically tagged” the resident
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