A transformation system for the yeast Candida utilis: use of a modified endogenous ribosomal protein gene as a drug-resistant marker and ribosomal DNA as an integration target for vector DNA

Abstract

We have developed a transformation system for the yeast Candida utilis. A novel strategy was applied to construct the transformation system, since auxotrophic mutants which could be used as hosts for transformation are not available. A gene encoding the ribosomal protein L41 was cloned from C. utilis, which is sensitive to cycloheximide, and used as a marker gene conferring cycloheximide resistance after modification of its amino acid sequence.

Efficient Gene Disruption in the High-Ploidy YeastCandida utilisUsing the Cre-loxPSystem

“…Furthermore, effective methods of transforming C. utilis have been developed, using electroporation and several bacterial antibiotic resistance markers, such as genes conferring resistance to G418 (APT), hygromycin B (HPT), or cycloheximide (mutated RPL41). 2,3) These techniques have since been used in the heterologous production of, e.g., monellin, -amylase, and carotenoids such as lycopene. [4][5][6][7] Compared with S. cerevisiae, however, the utilization of genetically modified C. utilis has been hampered due to its polyploidy, its imperfect life cycle, and the limited set of selection-marker genes.…”

Genetic Engineering ofCandida utilisYeast for Efficient Production ofL-Lactic Acid

“…Construction of a C. utilis genomic DNA library has been described. 22) In order to obtain fragments containing the PDC gene, PCR was performed using C. utilis genomic DNA as template. Oligonucleotides IKSM-29 and IKSM-30 were used as primers.…”

“…[18][19][20] C. utilis can grow on inexpensive substrates, e.g., pulping-waste liquors from the paper industry, while most other yeasts cannot. 21) Since the development of an efficient electroportation-based method for transforming C. utilis, 22) this yeast has been used in the heterologous production of monellin, y To whom correspondence should be addressed. Fax: +81-45-788-4042; E-mail: Shigehito Ikushima@kirin.co.jp Abbreviations: kb, kilo-base pair; HygB, hygromycin B; PDC, pyruvate decarboxylase; L-LDH , L-lactate dehydrogenase; AcH, acetaldehyde Biosci.…”

Ethanol Production from Xylose by a RecombinantCandida utilisStrain Expressing Protein-Engineered Xylose Reductase and Xylitol Dehydrogenase

“…[20][21][22] Since the development of an efficient electroporation-based method for transforming C. utilis, 23) this yeast has been used for the heterologous production of monellin, 24) -amylase, 25) and carotenoids such as lycopene. [26][27][28] Recently, we developed efficient transformation systems for C. utilis, 29) and to our knowledge achieved the highest level of production of L-lactic acid by a yeast species using a geneticallyengineered strain of C. utilis.…”