DEVELOPMENT OF MHV-RESISTANT MOUSE USING RNAi TRAP METHOD

动植物体内转录后基因沉默称为RNA干扰(RNAi)。RNAi是双链RNA(DsRNA)诱导同源mRNA序列依赖的降解过程。RNAi的天然作用可能诱导对病毒感染的防御和对细胞基因表达的调节。遗传和生物学研究表明，RNAi是一个非常复杂的过程，涉及许多不同的蛋白质，功能大多未知。近年来的研究表明，21-核苷酸(NT)合成的小干扰RNA(SiRNAs)和由U6或H1启动子转录的siRNA能特异性地抑制多种哺乳动物细胞内源性基因的表达。这些21个核苷酸的siRNA双链能够逃避INF防御系统，这些发现表明哺乳动物中可能存在RNAi或与RNAi相关的系统。小鼠肝炎病毒(MHV)感染是当代实验群体中的一种重要病毒感染。最近，转基因和基因敲除小鼠频繁地在研究人员之间交换，导致各国MHV污染的高发。目前，我们没有有效的预防感染的方法，也没有任何治疗方法。预防感染的一种方法是培育出一只耐MHV的小鼠。为此，我们尝试建立了一种RNAi基因捕获方法。我们已成功地建立了一种可互换的基因捕获方法。使用这种方法，我们可以在小鼠身上进行大规模的诱变。如果能同时培育出对MHV有抵抗力的基因陷阱小鼠，将对这些小鼠的分布有很大的帮助。然而，我们开发的方法仍然不能有效地阻止RNA基因的表达。要达到最初的目的，还需要进一步的研究。

The post-transriptional gene silencing in animals and plants is called RNA interference (RNAi). RNAi is a process in that double-stranded RNA (dsRNA) induces a sequence-dependent degradation of a cognate mRNA. The natural roles of RNAi might induce defense against viral infection and regulation of the expression of cellular genes. Genetic and biological studies revealed that RNAi is a very complex process that involves many different proteins with mostly unidentified functions. It was reported recendy that 21-nucleotide (nt) synthetic small interfering RNAs (siRNAs) and siRNA that were transcribed by U6 or H1 promoter specifically suppressed the expression of endogenous genes in several lines of mammalian cells. These 21-nt siRNA duplexes were able to evade the INF defense system and these findings suggested that RNAi or an RNAi-related system might exist in mammals.Mouse hepatitis virus (MHV) infection is an important viral infection in contemporary laboratory colonies. Recently, transgenic and knockout mice are frequently exchanged between investigators leading to the high incidence of contamination of MHV across the countries. At present, we do not have effective methods to prevent infection, nor do we have any treatment. One way to prevent infection is to produce a MHV-resistant mouse. For this purpose, we tried to develop an RNAi gene trap method. We previouly succeeded to develop an exchangeable gene trap method. Using this method, we can carry out a large scale mutagenesis in mice. If we can produce a gene trap mouse which is resistant to MHV at the same time, it will be quite helpful to distribute these mice. However, the method we developed is not still effective to prevent RNA gene expression. Further studies will be required to accomplish the initial purpose.