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RNA interference as a metabolic engineering tool: potential for in vivo control of protein expression in an insect larval model.
Metabolic Engineering ( IF 8.4 ) Pub Date : 2003-09-02 , DOI: 10.1016/s1096-7176(03)00027-2
Shannon F Kramer 1 , William E Bentley
Affiliation  

Many ex vivo factors influence the yield of recombinant protein produced via AcMNPV (Autographa californica multiple nucleocapsid nuclear polyhedrosis virus) in Trichoplusia ni (T. ni) larvae. Among these are: the method of infection, the time of infection, the virus load, and the time of harvest. In vivo strategies, however, that attempt to manipulate host function in this and other expression systems have largely been ignored. In this work, RNA interference (RNAi) is shown as an effective metabolic engineering controller to downregulate targeted gene expression. Specifically, RNAi was made to virus-encoded gfp(uv) and was found to inhibit the production of GFPuv in larvae when injected within an 18-h window (before and after) of baculovirus infection. The level of inhibition was found to depend, both in duration and extent, on the concentration of injected RNAi. That relatively low levels of RNAi can inhibit protein synthesis driven by the strong polyhedrin (polh) promoter of AcMNPV, suggests that RNAi will find utility as an in vivo metabolic controller in metabolic engineering studies such as this one pertaining to protein expression.

中文翻译:

RNA干扰作为代谢工程工具:在昆虫幼虫模型中体内控制蛋白质表达的潜力。

许多离体因素会影响粉虱(Trichoplusia ni(T. ni))幼虫中通过AcMNPV(加利福尼亚州产自噬菌种多核衣壳核多角体病毒)产生的重组蛋白的产量。其中包括:感染方法,感染时间,病毒载量和收获时间。然而,在体内策略中试图操纵该表达系统和其他表达系统中的宿主功能的尝试已被大大忽略。在这项工作中,RNA干扰(RNAi)被显示为有效下调靶向基因表达的代谢工程控制者。具体而言,将RNAi制成病毒编码的gfp(uv),并发现当在杆状病毒感染的18小时窗内(前后)注射时,RNAi可抑制幼虫中GFPuv的产生。发现抑制水平取决于持续时间和程度,注射RNAi的浓度。RNAi的相对较低水平可以抑制由AcMNPV的强多角体蛋白(polh)启动子驱动的蛋白质合成,这表明RNAi将在代谢工程研究(例如与蛋白质表达有关的研究)中用作体内代谢控制物。
更新日期:2019-11-01
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