A site-directed integration system for the nonuniversal CUG(Ser) codon usage species Pichia farinosa by electroporation.
Halotolerant yeast, Pichia farinosa, is a valuable yeast strain in fermentation industry because it produces high yield of glycerol and xylitol, and can tolerate both contamination and high-density growth during fermentation. However, the lack of genetic manipulation tools makes it less popular as a gene engineering strain. Expression systems commonly used in other yeast systems, such as Saccharomyces cerevisiae and Pichia pastoris cannot be used in P. farinosa because it translates universal Leu codon CUG as Ser. Here we reported a modified expression vector and a transformation system with enhanced efficiency in P. farinosa. The results showed that cells of OD(600 )0.8-1.0 with DTT treatment can obtain high transformation efficiency. The optimized electroporation condition was 900 V, 25 microF, and 200 Omega. The DNA concentration did not influence the transformation. Our system provides the potential not only for applying P. farinosa as an industrial strain of gene engineering, but also for studying gene function in its native host.
Wang
Xiaoxing
X
Department of Biology, Sichuan University, Wangjiang Street No.29, Chengdu, Sichuan, China.
Li
Gang
G
Deng
Yuntao
Y
Yu
Xuanwei
X
Chen
Fang
F
D000595:Amino Acid Sequence; D001483:Base Sequence; D001761:Bleomycin / Q000494:pharmacology; D003062:Codon / Q000235:genetics*; D018274:Electroporation / Q000379:methods*; D005818:Genetic Engineering / Q000379:methods; D005822:Genetic Vectors; D005839:Gentamicins / Q000494:pharmacology; D008969:Molecular Sequence Data; D010843:Pichia / Q000187:drug effects / Q000235:genetics* / Q000254:growth & development; D014170:Transformation, Genetic
D003062:Codon; D005839:Gentamicins; D001761:Bleomycin; C105427:Zeocin; C010680:antibiotic G 418