[含禽冠状病毒免疫原性基因的转基因马铃薯及其小鼠免疫原性]。

Jian-Xiang Wu, Ji-Yong Zhou, Xue-Ping Zhou
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引用次数: 0

摘要

为验证禽冠状病毒传染性支气管炎病毒(IBV)免疫原性基因在植物中表达的可行性,研究了IBV S1基因在马铃薯中的转化及其表达产物的免疫原性。在35s启动子控制下,将IBV-ZJ971菌株S1基因插入到pBI121质粒中。用三亲本杂交方法获得了重组载体pBI121的产烟农杆菌EHA105。以此为基础,建立了一种由制烟农杆菌介导的马铃薯高效转化体系。转IBV S1基因马铃薯愈伤组织分化率为100%,芽分化率为95%以上。PCR和Southern blot分析表明,IBV S1基因被整合到马铃薯植株的基因组DNA中,大多数转基因植株具有IBV S1基因的两个拷贝。在我们的实验中,获得了47个转基因植株。Northern blot和ELISA分析表明,大多数转基因植株都能正常转录和翻译IBV S1基因,但不同转基因植株的转录和翻译水平不同。BALB/C小鼠免疫试验表明,含IBV S1基因的转基因马铃薯表达产物具有免疫原性,在0.5 g和1 g剂量下,ELISA抗体效价分别达到1:20 ~ 1:40和1:80 ~ 1:160。采用气管器官培养法检测病毒中和抗体(VN),结果表明,在0.5 g和1 g剂量下,VN滴度分别为1:160 ~ 1:20 20和1:20 20 ~ 1:2048。
本文章由计算机程序翻译,如有差异,请以英文原文为准。
[Transgenic potato containing immunogenic gene of avian coronavirus and its immunogenicity in mice].

To check the feasibility of expression of the immunogenic gene of avian coronavirus infectious bronchitis virus (IBV) in plants, the transformation of S1 gene of IBV into potato and the immunogenicity of its expression product was studied. The S1 gene of IBV-ZJ971 strain was inserted into plasmid pBI121 under the control of 35 S promoter. Agrobacterium fumefaciens EHA105 with the recombinant vector pBI121 was obtained by tri-parental mating method. So, an efficient potato transformation system mediated by Agrobacterium fumefaciens was established. The rates of calli and shoots differentiation were 100%, and more than 95% respectively, for transgenic potato with S1 gene of IBV. PCR and Southern blot analyses showed that IBV S1 gene was integrated into genomic DNA of the potato plant and most transgenic plants had two copies of S1 gene of IBV. In our experiments, 47 transgenic plantlets have been obtained. Northern blot and ELISA analyses indicated that most transgenic plants could normally transcribe and translate S1 gene of IBV, though the levels of transcription and translation were different in various transgenic plants. Immunity assay with BALB/C mice showed that expression products of transgenic potato with S1 gene of IBV were immunogenic, and ELISA antibody titer reached 1:20 to 1:40 and 1:80 to 1:160 with doses of 0.5 g and 1 g, respectively. Virus neutralization (VN) antibodies were detected by tracheal organ cultures, and the results showed that VN titers reached respectively 1:160 to 1:320 and 1:320 to 1:2048 with doses of 0.5 g and 1 g.

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