在9月12日的《美國科學(xué)院院刊》上，來自美國加州理工學(xué)院和加州大學(xué)Veterans Affairs醫(yī)學(xué)中心的研究人員描述了一種新的慢病毒(lentiviral)載體平臺pSLIK（用于誘導(dǎo)敲除的單慢病毒載體）,，這種載體能使研究人員通過對任何細(xì)胞系統(tǒng)進(jìn)行單個的病毒感染,，并用四環(huán)素調(diào)節(jié)類microRNA短發(fā)夾RNA的表達(dá),。
越來越多的證據(jù)表明RNAi技術(shù)是對哺乳動物基因組功能分析的一種強(qiáng)大的實驗工具。研究人員通過尋找能夠使內(nèi)源基因表達(dá)調(diào)節(jié)與外源轉(zhuǎn)入基因的再次協(xié)同表達(dá)的方法將會揭示出這種技術(shù)的全部潛能,。
在PNAS上的這篇新文章中,，研究人員描述了一個新的慢病毒載體平臺pSLIK的研發(fā)過程。
在小鼠胚胎纖維原細(xì)胞中,，pSLIK平臺能夠被用來選擇性地抑制heterotrimeric G蛋白G 12 和/或 G 13的表達(dá),，并且證實G 13的轉(zhuǎn)錄依賴血清應(yīng)答成分。
在RAW264.7巨噬細(xì)胞中,，G 2的調(diào)節(jié)性敲除與鈣離子對C5a的反應(yīng)下降相一致,。在G 2的類microRNA短發(fā)夾RNA上游插入GFP（綠色熒光蛋白基因）基因能使一個異源mRNA在利用四環(huán)素進(jìn)行靶向基因敲除時發(fā)生再次表達(dá)。這種做法明顯提高了pSLIK系統(tǒng)的實際操作的可行性,。
部分英文原文：
A single lentiviral vector platform for microRNA-based conditional RNA interference and coordinated transgene expression
RNAi is proving to be a powerful experimental tool for the functional annotation of mammalian genomes. The full potential of this technology will be realized through development of approaches permitting regulated manipulation of endogenous gene expression with coordinated reexpression of exogenous transgenes. We describe the development of a lentiviral vector platform, pSLIK (single lentivector for inducible knockdown), which permits tetracycline-regulated expression of microRNA-like short hairpin RNAs from a single viral infection of any naïve cell system. In mouse embryonic fibroblasts, the pSLIK platform was used to conditionally deplete the expression of the heterotrimeric G proteins G12 and G13 both singly and in combination, demonstrating the G13 dependence of serum response element-mediated transcription. In RAW264.7 macrophages, regulated knockdown of G2 correlated with a reduced Ca2+ response to C5a. Insertion of a GFP transgene upstream of the G2 microRNA-like short hairpin RNA allowed concomitant reexpression of a heterologous mRNA during tetracycline-dependent target gene knockdown, significantly enhancing the experimental applicability of the pSLIK system.
