Kousaku Murata

Intact yeast cells treated with alkali cations took up plasmid DNA. Li+, Cs+, Rb+, K+, and Na+ were effective in inducing competence. Conditions for the transformation of Saccharomyces cerevisiae D13-1A with plasmid YRp7 were studied in detail with CsCl. The optimum incubation time was 1 h, and the optimum cell concentration was 5 x 10(7) cells per ml. The optimum concentration of Cs+ was 1.0 M. Transformation efficiency increased with increasing concentrations of plasmid DNA. Polyethylene glycol was absolutely required. Heat pulse and various polyamines or basic proteins stimulated the uptake of plasmid DNA. Besides circular DNA, linear plasmid DNA was also taken up by Cs+-treated yeast cells, although the uptake efficiency was considerably reduced. The transformation efficiency with Cs+ or Li+ was comparable with that of conventional protoplast methods for a plasmid containing ars1, although not for plasmids containing a 2 microns origin replication.

Transformation (i.e., genetic modification of a cell by the incorporation of exogenous DNA) is indispensable for manipulating fungi. Here, we review the transformation methods for Saccharomyces cerevisiae, Schizosaccharomyces pombe, Candida albicans, Pichia pastoris and Aspergillus species and discuss some common modifications to improve transformation efficiency. We also present a model of the mechanism underlying S. cerevisiae transformation, based on recent reports and the mechanism of transfection in mammalian systems. This model predicts that DNA attaches to the cell wall and enters the cell via endocytotic membrane invagination, although how DNA reaches the nucleus is unknown. Polyethylene glycol is indispensable for successful transformation of intact cells and the attachment of DNA and also possibly acts on the membrane to increase the transformation efficiency. Both lithium acetate and heat shock, which enhance the transformation efficiency of intact cells but not that of spheroplasts, probably help DNA to pass through the cell wall.

Bioethanol production from algae is a promising approach that resolves problems associated with biofuel production from land biomass, such as bioethanol–food conflicts and the indirect land use change. However, it presents several technical difficulties because existing ethanologenic microbes can neither degrade alginate, a major component of brown algae, nor assimilate alginate degradation products. We developed an integrated bacterial system for converting alginate to ethanol using a metabolically modified, alginate-assimilating, pit-forming bacterium, Sphingomonas sp. A1 (strain A1). Overexpression of Zymomonas mobilis pdc and adhB was achieved using a strong constitutive expression promoter newly identified in strain A1 and by inserting multiple gene copies using the methylation sensitivity of XbaI. Metabolome analysis revealed by-product accumulation, and its synthesis pathway was blocked by gene disruption. The ethanologenic recombinant strain A1 accumulated 13.0 g L−1ethanol in 3 d using alginate as the sole carbon source.

Journal Article Effect of Depolymerized Alginates on the Growth of Bifidobacteria Get access Hisayoshi Akiyama, Hisayoshi Akiyama Otsuka Chemical Co., Ltd., Division of Food and Beverage Research, Kawauchi, Tokushima 771–01, Japan Search for other works by this author on: Oxford Academic Google Scholar Taeko Endo, Taeko Endo Otsuka Chemical Co., Ltd., Division of Food and Beverage Research, Kawauchi, Tokushima 771–01, Japan Search for other works by this author on: Oxford Academic Google Scholar Ryo Nakakita, Ryo Nakakita Otsuka Chemical Co., Ltd., Division of Food and Beverage Research, Kawauchi, Tokushima 771–01, Japan Search for other works by this author on: Oxford Academic Google Scholar Kousaku Murata, Kousaku Murata Otsuka Chemical Co., Ltd., Division of Food and Beverage Research, Kawauchi, Tokushima 771–01, JapanResearch Institutefor Food Science, Kyoto University, Uji, Kyoto 611, Japan Corresponding author. Search for other works by this author on: Oxford Academic Google Scholar Yoshimasa Yonemoto, Yoshimasa Yonemoto Otsuka Chemical Co., Ltd., Division of Food and Beverage Research, Kawauchi, Tokushima 771–01, Japan Search for other works by this author on: Oxford Academic Google Scholar Kenichi Okayama Kenichi Okayama Otsuka Chemical Co., Ltd., Division of Food and Beverage Research, Kawauchi, Tokushima 771–01, Japan Search for other works by this author on: Oxford Academic Google Scholar Bioscience, Biotechnology, and Biochemistry, Volume 56, Issue 2, 1 January 1992, Pages 355–356, https://doi.org/10.1271/bbb.56.355 Published: 01 January 1992 Article history Received: 13 August 1991 Published: 01 January 1992