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Title: TRANSFORMATION OF ALTERNAN STRAINS OF LEUCONOSTOC BY ELECTROPORATION

Author
item LEATHERS, TIMOTHY
item JONES, JAMES - NORTHWESTERN UNIV
item WYCKOFF, HERB - CREST FOODS

Submitted to: Biotechnology Letters
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 5/11/2004
Publication Date: 7/1/2004
Citation: Leathers, T.D., Jones, J.D., Wyckoff, H.A. 2004. Transformation of alternan-producing strains of Leuconostoc by electroporation. Biotechnology Letters. 26(14):1119-1124.

Interpretive Summary: Value-added food and nonfood products and processes are needed to enhance the economic viability and competitiveness of U.S. agriculture. Novel microbial gums represent potential new value-added products as well as domestic replacements for imported plant gums. However, new genetic tools are needed to understand and manipulate the production of these gums. This work will be of interest to researchers developing new uses and value-added products from agricultural commodities and byproducts.

Technical Abstract: Alternan-producing Leuconostoc mesenteroides strain NRRL B-1355 and its glucansucrase-negative derivative NRRL B-21414 were transformed by electroporation using four gram positive-gram negative shuttle vectors. Optimal conditions were 400 ohms and 10 kV/cm, resulting in transformation frequencies of up to 3.5 x 10**4 per microgram DNA. Relatively low copy numbers and native plasmids made it difficult to visualize the introduced plasmids on ethidium bromide-strained gels and in some cases on blot hybridizations. However, PCR analysis indicated that 95% of putative transformants carried plasmid sequences. Direct colony PCR was shown to work well for this system and also for transformants of L. mesenteroides subsp. cremoris.