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禽呼肠孤病毒σC蛋白在转基因植物中的表达。

Expression of avian reovirus sigmaC protein in transgenic plants.

作者信息

Huang Liang-Kai, Liao Sin-Chung, Chang Ching-Chun, Liu Hung-Jen

机构信息

Institute of Biotechnology, National Cheng Kung University, Tainan 701, Taiwan.

出版信息

J Virol Methods. 2006 Jun;134(1-2):217-22. doi: 10.1016/j.jviromet.2006.01.013. Epub 2006 Feb 20.

Abstract

Avian reovirus (ARV) structural protein, sigmaC encoded by S1 genome segment, is the prime candidate to become a vaccine against ARV infection. Two plant nuclear expression vectors with expression of sigmaC-encoding gene driven by CaMV 35S promoter and rice actin promoter were constructed, respectively. Agrobacterium containing the S1 expression constructs were used to transform alfalfa, and transformants were selected using hygromysin. The integration of S1 transgene in alfalfa chromosome was confirmed by PCR and histochemical GUS staining. Western blot analysis using antiserum against sigmaC was carried out to determine the expression of sigmaC protein in transgenic alfalfa cells. The highest expression levels of sigmaC protein in the cellular extracts of selected p35S-S1 and pAct1-S1 transgenic alfalfa lines were 0.008% and 0.007% of the total soluble protein, respectively. The transgenic alfalfa cells with expression of sigmaC protein pave the way for the development of edible vaccine.

摘要

禽呼肠孤病毒(ARV)的结构蛋白σC由S1基因组片段编码,是开发抗ARV感染疫苗的主要候选蛋白。分别构建了两种植物核表达载体,其编码σC的基因表达由花椰菜花叶病毒(CaMV)35S启动子和水稻肌动蛋白启动子驱动。将含有S1表达构建体的农杆菌用于转化苜蓿,并使用潮霉素筛选转化体。通过PCR和组织化学GUS染色确认了S1转基因在苜蓿染色体中的整合。使用抗σC抗血清进行蛋白质免疫印迹分析,以确定σC蛋白在转基因苜蓿细胞中的表达。在选定的p35S-S1和pAct1-S1转基因苜蓿系的细胞提取物中,σC蛋白的最高表达水平分别占总可溶性蛋白的0.008%和0.007%。表达σC蛋白的转基因苜蓿细胞为食用疫苗的开发铺平了道路。

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