Intracellular release of recombinant green fluorescent protein (gfp(uv)) from Escherichia coli.

Thereza Christina Vessoni Penna, Eb Chiarini, Irene Alexeevna Machoshvili, Marina Ishii, Adalberto Pessoa

Department of Biochemical and Pharmaceutical Technology, School of Pharmaceutical Science, University of São Paulo, SP, Brazil. tcvpenna@usp.br

Applied biochemistry and biotechnology 2002

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The recombinant green fluorescent protein (gfp(uv)) was expressed by Escherichia coli DH5-alpha cells transformed with the plasmid pGFPuv. The gfp(uv) was selectively permeabilized from the cells in buffer solution (25 mM Tris-HCl, pH 8.0), after freezing (-70 degrees C for 15 h), by four freeze (-20 degrees C)/thaw cycles interlaid by sonication. The average content of released gfp(uv) (experiment 2) was 7.76, 34.58, 39.38, 12.90, and 5.38%, for the initial freezing (-70 degrees C) and the first, second, third and fourth freeze/thaw cycles, respectively. Superfusion on freezing was observed between -11 degrees C and -14 degrees C, after which it reached -20 degrees C at 0.83 degrees C/min.

Citation

Thereza Christina Vessoni Penna, Eb Chiarini, Irene Alexeevna Machoshvili, Marina Ishii, Adalberto Pessoa. Intracellular release of recombinant green fluorescent protein (gfp(uv)) from Escherichia coli. Applied biochemistry and biotechnology. 2002;98-100:791-802