【摘要】：目的:構(gòu)建川澤瀉鯊烯合酶(squalene synthase,SS)基因的原核表達載體。方法:在建澤瀉SS基因序列的基礎(chǔ)上,采用RT-PCR技術(shù),從川澤瀉新鮮葉片中克隆得到SS基因,并在BL21(DE3)細(xì)胞中分別用不同的表達載體、IPTG濃度、溫度誘導(dǎo)該蛋白表達。結(jié)果:該基因在大腸桿菌中表達,但為包涵體,將其克隆到帶有GST標(biāo)簽的pGEX-6t-1載體中進行原核表達并未改善蛋白溶解性;30℃誘導(dǎo)蛋白表達最佳;不同IPTG濃度對蛋白表達影響不大。結(jié)論:川澤瀉的原核表達為后續(xù)對該基因的生物學(xué)功能及川澤瀉品質(zhì)改良奠定了基礎(chǔ)。
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圖片說明：川澤瀉總ＲNA凝膠電泳圖
[Abstract]:Objective: to construct the prokaryotic expression vector of squalene polymerase (squalene synthase,SS) gene in Radix et Rhizoma Chuanze. Methods: on the basis of SS gene sequence, SS gene was cloned from fresh leaves of alisma orientalis by RT-PCR technique, and the expression of SS gene was induced by different expression vectors, IPTG concentration and temperature in BL21 (DE3) cells. Results: the gene was expressed in E. coli, but it was an inclusion body. Cloning it into pGEX-6t-1 vector with GST tag for prokaryotic expression did not improve the solubility of the protein. The best protein expression was induced at 30 鈩，

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