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ARS Home » Research » Publications at this Location » Publication #70435
Title: IMPROVED ELECTROPORATION PROTOCOL AND VECTORS FOR STREPTOCOCCUS BOVIS

Author
item Wyckoff, Herbert
item WHITEHEAD, TERENCE

Submitted to: World Journal of Microbiology and Biotechnology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 6/19/1996
Publication Date: N/A
Citation: N/A

Interpretive Summary: New and efficient bioprocesses are needed for the conversion of agricultural commodities into biofuels and biopolymers. One research approach is to use a bacterium that is capable of growing directly on corn starch without the use of supplemental enzymes and genetically engineer the bacterium to produce fuel ethanol. We now report an improved method for introduction of genetic material into a rumen bacterium capable of excellent growth on corn starch. This method, along with improved genetic delivery systems, should allow researchers to genetically modify this bacterium for production of ethanol and other value-added products.

Technical Abstract: An improved method for electroporation of the ruminal bacterium Streptococcus bovis was developed. The organism was grown aerobically in hyperosmotic medium in the presence of glycine, and electroporation was carried out in a sucrose-glycerol solution with a field strength of 12.5 kV cm**-1, 200 ohms resistance and 25 uF capacitance. Routine electroporation efficiencies of 0.5-2.0 X 10**5 transformants/ug DNA were achieved. Improved vectors for S. bovis were developed that include a multiple cloning site and also a promoter region from the S. bovis intracellular amylase gene that may serve as an expression system for foreign genes.