CAJ | 학술논문

用PCR方法克隆了1.5kb的酿酒酵母Sacchromyces cerevisiae 海藻糖合成酶基因TPS1，将该片段连接到pUC19载体，通过转化分别引入海藻糖合成酶基因缺失和缺陷的大肠杆菌Escherichia coli FF4169和FF4050。对转化株的质粒DNA酶切分析表明均含有1.5kbPCR克隆片段。生长曲线实验证明，带有克隆片段的转化株在含0.5mol/L NaCl的高渗透压基础培养基中生长良好；用高效液相色谱(HPLC)结合蒸发光散射(ELSD)技术测定细胞内海藻糖实验证明转化株能够合成海藻糖。
용PCR방법극륭료1.5kb적양주효모Sacchromyces cerevisiae 해조당합성매기인TPS1，장해편단련접도pUC19재체，통과전화분별인입해조당합성매기인결실화결함적대장간균Escherichia coli FF4169화FF4050。대전화주적질립DNA매절분석표명균함유1.5kbPCR극륭편단。생장곡선실험증명，대유극륭편단적전화주재함0.5mol/L NaCl적고삼투압기출배양기중생장량호；용고효액상색보(HPLC)결합증발광산사(ELSD)기술측정세포내해조당실험증명전화주능구합성해조당。
A S. cerevisiae TPS1 gene for trehalose-6-phosphate synthase was cloned by PCR amplification. The 1.5kb DNA fragment was ligated to pUC19 and transformed into otsA deficient and deleted of E. coli strains FF4169 and FF4050 separately. otsA gene is encoding trehalose-6-phasphate synthase in E.coli. Restriction endonucleases digestion analysis of transformants＇ plasmid DNA showed that there was a cloned 1.5kb fragment carried on the vector. The growth curve experiment result showed that the both transformants could grow well as wild type. Trehalose was synthesized and accumulated in these transformants during high osmotic stress by HPLC conbined with ELSD (Evaporate light scatter detactor) determination. From the result above, we could conclude that the TPS1 gene of S.cerevisiae was able to restore otsA gene function in E.coli for both osmotolerance and trehalose accumulation during salt stress.