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題 名 | 黑鯛第一型類胰島素生長因子之融合蛋白質溶解性的研究=The Study of the Solubility of Black Seabream Insulin-Like Growth Factor I Fusion Protein |
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作 者 | 吳鴻程; 朱惠鈴; 張繼堯; 吳金洌; | 書刊名 | 中國農業化學會誌 |
卷 期 | 36:6 1998.12[民87.12] |
頁 次 | 頁612-621 |
分類號 | 387.1015 |
關鍵詞 | 第一型類胰島素生長因子; 質體pGEX-3X; 蛋白質溶解性; 黑鯛; 界面活性劑; 培養溫度; Insulin-like growth factor I; Plasmid pGEX-3X; Solubilization; Black seabream; Sarkosyl; Culture temperature; |
語 文 | 中文(Chinese) |
中文摘要 | 本研究以核��聚合�t連鎖反應,合成黑鯛第一型類胰島生長因子(insulin-like growth factor 1, IGF-I)基因,再接合到表現質體pGEX-3X中,成為質體pGEX3X-IGF-I。 質體轉型到大腸桿菌,經isopropyl β-D-thiogalactopyranoside(IPTG)誘發,可大量表 現生成GST(glutathions S-transferase)和IGF-I融合的GST-IGF-I融合蛋白質,其分子量 為34.5KD。質體的宿主菌株Xa90,TG1及BL21(DE3)中,以BL21(DE3)生成融合蛋白質較 多。使宿主菌體於30 ℃培養及添加1.3% sarkosyl 於振破液中,有助於水溶性融合蛋白質 的生成及抽取;減短IPTG誘發時間,無法促進菌體生成水溶性融合蛋白質。 |
英文摘要 | A pGEX3X-IGF-I recombinant plasmid was constructed by cloning a PCR product of the IGF-I(insulin-like growth factor I)gene of Black Seabream into the pGEX-3X expression vector. The tac promoter of pGEX3X-IGF-I directed the synthesis of IGF-I as a fusion protein with the C terminal of GST(glutathione S- transferase). Induction of the tac promoter with isopyropyl β-D- thiogalactopyranos-ide (IPTG)in E. coli transformed with pGEX3X-IGF-I resulted in the over-expression of a 34.5 KD GST-IGF-I fusion protein. When this plasmid is transformed into different E. coli hosts, we found that BL21(DE3) had the maximum expression, followed by Xa90 and TG1. The solubility of fusion protein could be improved by incubating the cell at 30℃. The extractive efficency could also be 3 days' of infection, respectively. Reduced body weight was also found in the surviving infected larvae, and treatment with four times dilution of the extract resulted in the body weight being less than one-tenth of normal larvae. Among the fractions separated by a silica gel column of silica gel, the fractions containing 32%DA, 31%DB, and 4.5%DE had equal virulence to the original extract. To each single DTX, the DE had the highest activity with 43.34% mortality and followed in order by DA 18.33%, DA��11.86%, DD 10%, and DB 2.27%. |
本系統中英文摘要資訊取自各篇刊載內容。