Translational enhancement of foreign protein expression by HSP101

• Main Authors: Pin-sheng Huang, 黃品升
• Other Authors: Ching-Chun Chang
• Format: Others
• Language: zh-TW
• Published: 2007
• Online Access: http://ndltd.ncl.edu.tw/handle/58977354498767854051

碩士 === 國立成功大學 === 生物科技研究所碩博士班 === 95 === Transgenic plants have been generated as bioreactors to produce proteins. The advantages of plant bioreactors include low-cost, easily and extensively cultivation. Edible vaccine, antibodies, and the therapeutical or industrial proteins, all of them have been produced by plant genetic engineering. However, the protein expression level in transgenic plant are low, and this is the reason to limit the development of plant as bioreactors. In previous study, the 68 base of TMV 5’-UTR called the Ω sequence, is able to promote the translational efficiency. The plant HSP101 can bind to the Ω sequence and recruit the translational initiation factors to enhance the translational activity of RNA transcripts. In this study we investigated the HSP101 overexpression lines in regulation of Ω-reporter gene expression in planta as well as in protoplast. In protoplast aspect, we used the electroporation method to deliver the Ω sequence and luciferase gene to the tobacco protoplasts and study the relationship of reporter gene (luciferase) expression level with HSP101. In our study, Ω sequence enhance luciferase expression level about 7~12 fold in OsHSP101 overexpressed tobacco protoplasts. In planta aspect, we had transformed tobacco with Ω-luciferase expression cassette by Agrobacterium-mediated transformation. Tree Ω-luciferase transgenic tobacco lines were obtained. We crossed the Ω-luciferase transgenic plants with HSP101 overexpression lines or wild type plants, and analyzed luciferase activity in the progenies. The results shown that OsHSP101 can enhance the reporter gene expression with Ω sequence in the 5’-UTR.