원문정보
초록
영어
Kluyveromyces marxianus is now considered as one of the best choice of option for industrial applications of yeast because the strain is able to grow at high temperature, utilizes various carbon sources and grows fast. In spite of these great advantages over other yeast strains, the use of K. marxianus as a host for industrial applications is still limited. This limitation results in part from lack of genetic engineering tools compared to its sister yeast strains such as Saccharomyces cerevisiae, and in part from lack of precise knowledge about the characteristics of currently-available molecular tools. Among them, characteristic of promoters is of prior importance because they control the time and the amount of protein expression. In this study, 4 well-known constitutive promoters (CYC, TEF, GPD and ADH) of S. cerevisiae were characterized in K. marxianus in terms of protein expression level and their stochastic behavior. For this purpose, 5 Ura-auxtrophic K. marxianus strains were generated, and expression vectors were constructed by inserting autonomous replicating sequence (ARS) and centromere sequences (CEN) of K. marxianus KCTC 17555 strain into pRS306 vector. Then, four cassettes comprising one of each promoter- green fluorescence protein (GFP)-TEF terminator were inserted into the vector. GFP expression under the control of one of each promoter was analyzed by reverse transcription PCR, fluorescence microscopy and flow cytometer. Promoter strength was in the order of GPD>ADH>TEF>>CYC. CYC ~30, TEF 200, 500, GPD 400, 900, ADH 100, 250Exclude plasmid lost. Exclude mitotic double expression