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首页> 美国卫生研究院文献>International Journal of Molecular Sciences >Enhancement of Recombinant Protein Production in Transgenic Nicotiana benthamiana Plant Cell Suspension Cultures with Co-Cultivation of Agrobacterium Containing Silencing Suppressors
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Enhancement of Recombinant Protein Production in Transgenic Nicotiana benthamiana Plant Cell Suspension Cultures with Co-Cultivation of Agrobacterium Containing Silencing Suppressors

机译：与含有沉默抑制剂的农杆菌共培养提高转基因烟草本氏烟草植物细胞悬浮培养中重组蛋白的生产

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We have previously demonstrated that the inducible plant viral vector (CMViva) in transgenic plant cell cultures can significantly improve the productivity of extracellular functional recombinant human alpha-1-antiryspin (rAAT) compared with either a common plant constitutive promoter (Cauliflower mosaic virus (CaMV) 35S) or a chemically inducible promoter (estrogen receptor-based XVE) system. For a transgenic plant host system, however, viral or transgene-induced post-transcriptional gene silencing (PTGS) has been identified as a host response mechanism that may dramatically reduce the expression of a foreign gene. Previous studies have suggested that viral gene silencing suppressors encoded by a virus can block or interfere with the pathways of transgene-induced PTGS in plant cells. In this study, the capability of nine different viral gene silencing suppressors were evaluated for improving the production of rAAT protein in transgenic plant cell cultures (CMViva, XVE or 35S system) using an Agrobacterium-mediated transient expression co-cultivation process in which transgenic plant cells and recombinant Agrobacterium carrying the viral gene silencing suppressor were grown together in suspension cultures. Through the co-cultivation process, the impacts of gene silencing suppressors on the rAAT production were elucidated, and promising gene silencing suppressors were identified. Furthermore, the combinations of gene silencing suppressors were optimized using design of experiments methodology. The results have shown that in transgenic CMViva cell cultures, the functional rAAT as a percentage of total soluble protein is increased 5.7 fold with the expression of P19, and 17.2 fold with the co-expression of CP, P19 and P24.
机译：我们先前已经证明，与任一常见植物组成型启动子（花椰菜花叶病毒（CaMV）相比，转基因植物细胞培养物中的可诱导植物病毒载体（CMViva）可以显着提高细胞外功能重组人α-1-antispin（rAAT）的生产率。 ）35S）或化学诱导型启动子（基于雌激素受体的XVE）系统。但是，对于转基因植物宿主系统，病毒或转基因诱导的转录后基因沉默（PTGS）已被鉴定为可能显着减少外源基因表达的宿主反应机制。先前的研究表明，病毒编码的病毒基因沉默抑制剂可以阻断或干扰植物细胞中转基因诱导的PTGS的途径。在这项研究中，评估了九种不同的病毒基因沉默抑制剂在农杆菌介导的瞬时表达共培养过程中提高转基因植物细胞培养（CMViva，XVE或35S系统）中rAAT蛋白产生的能力。细胞和携带病毒基因沉默抑制剂的重组农杆菌在悬浮培养物中一起生长。通过共培养过程，阐明了基因沉默抑制剂对rAAT产生的影响，并鉴定了有希望的基因沉默抑制剂。此外，使用实验方法设计优化了基因沉默抑制剂的组合。结果表明，在转基因CMViva细胞培养物中，功能性rAAT作为总可溶性蛋白的百分比随P19的表达增加了5.7倍，而与CP，P19和P24的共表达则增加了17.2倍。

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