臺灣博碩士論文加值系統

利用水稻白化幼苗所分離出的水稻第一族低分子量熱休克蛋白質基因， oshsp16.9，及自豌豆白化幼苗所分離出的豌豆冷溫誘導蛋白質基因，peaci11.8，為材料，將這兩股cDNA分別構築 (construct) 到pGEX2T表現載體 (expression vector) 中，並轉型 (transformation) 到大腸桿菌中表現。利用適當濃度isopropyl β-D-thiogalactopyranoside (IPTG) 誘導，促使大腸桿菌大量表現融合蛋白質 glutathione S-transferase (GST)、GST-oshsp16.9及GST-peaci11.8。目前已知48℃ 及4℃ 都會造成pGEX2T的轉殖細胞死亡，因此將大量表現蛋白質的大腸桿菌移入48℃或4℃中處理。在48℃處理之下，計算細胞的存活率，發現轉殖熱休克蛋白質基因之大腸桿菌具有顯著的耐熱性，但相對地，轉殖pGEX2T之細胞則無；另外發現轉殖冷溫誘導蛋白質基因的大腸桿菌，在高溫處理下仍具有部份耐熱性，其存活率約為轉殖熱休克蛋白質基因之細胞的一半。在4℃的處裡之下，轉殖冷溫誘導蛋白質基因之大腸桿菌細胞表現出不錯的耐冷性，而轉殖熱休克蛋白質基因的細胞也具有部份的耐冷性。因此，不論轉殖熱休克蛋白質基因或冷溫誘導蛋白質基因的大腸桿菌，其在高溫或低溫逆境下，均可提高細胞不同程度的耐熱性或耐冷性。

Orzya sativa heat-shock protein gene (Oshsp16.9) was isolated from etiolated rice seedlings and pea cold-induced protein gene (peaci11.8)was isolated from etiolated pea seedlings. We constructed the recombinant Oshsp16.9 and peaci11.8 to expression vector, pGEX2T, and transformed into Escherichia coli XL1-Blue cells. After isopropyl β-D-thiogalactopyranoside (IPTG) induction, the transformed E. coli cells overexpressed the fusion proteins, glutathione S-transferase (GST), GST-Oshsp16.9 and GST-peaci11.8. After overexpressing the fusion proteins, cells were transferred to 48℃ and 4℃, the conditions lethal to pGEX2T cells. Under 48℃ treatment, pGEX2T-oshsp16.9 resulted in significant thermotolerance is similar to the previous reports. There was an interesting finding that pGEX2T-peaci11.8 also increased thermotolerance, and the efficiency was between that of pGEX2T and pGEX2T-oshsp16.9 revealed. However, different results were obtained at 4℃ treatment. A new finding revealed that pGEX2T-peaci11.8 enhanced the cell survival efficiency. The difference of survival was more than 50% after 3 days treatment. In addition, pGEX2T-oshsp16.9 could not only have thermotolerance but also increase cold-tolerance. In conclusion, in the prokaryotic expression system, whether the heat-shock protein gene or cold-induced protein gene from plants could cause different levels of thermotolerance or cold-tolerance under different temperature stress.

中文摘要---------------------------------------------------1
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