Isolation of Small Noncoding RNAs from Human Serum

The analysis of RNA and its expression is a common feature in many laboratories. Of significance is the emergence of small RNAs like microRNAs, which are found in mammalian cells. These small RNAs are potent gene regulators controlling vital pathways such as growth, development and death and much interest has been directed at their expression in bodily fluids. This is due to their dysregulation in human diseases such as cancer and their potential application as serum biomarkers. However, the analysis of miRNA expression in serum may be problematic. In most cases the amount of serum is limiting and serum contains low amounts of total RNA, of which small RNAs only constitute 0.4-0.5%¹. Thus the isolation of sufficient amounts of quality RNA from serum is a major challenge to researchers today. In this technical paper, we demonstrate a method which uses only 400 µl of human serum to obtain sufficient RNA for either DNA arrays or qPCR analysis. The advantages of this method are its simplicity and ability to yield high quality RNA. It requires no specialized columns for purification of small RNAs and utilizes general reagents and hardware found in common laboratories. Our method utilizes a Phase Lock Gel to eliminate phenol contamination while at the same time yielding high quality RNA. We also introduce an additional step to further remove all contaminants during the isolation step. This protocol is very effective in isolating yields of total RNA of up to 100 ng/µl from serum but can also be adapted for other biological tissues.     

新型非编码小RNA——Piwi-interacting RNA(piRNA)

piRNA(Piwi-interacting RNA)是最近从哺乳动物睾丸组织中发现的一类能与PIWI蛋白质相互作用,且长度分布在26～31nt的新型小分子单链RNA,主要综述piRNA的相关研究进展.     

A Versatile RNA Vector for Delivery of Coding and Noncoding RNAs

The discovery that RNA viruses, lacking any DNA intermediate, can be engineered to express both coding and noncoding RNAs suggests that this platform may have therapeutic value as a delivery vehicle. Here we illustrate that a self-replicating, noninfectious RNA, modeled on influenza virus, provides one such example of a versatile in vivo delivery system for silencing and/or expressing a desired RNA for therapeutic purposes.     

细菌非编码小RNA分子RyhB的结构与功能

RyhB是一种大小为90个核苷酸的细菌非编码小RNA分子(small noncoding RNA, sRNA).当铁缺乏时,RyhB通过下调一系列与铁的储存和利用相关蛋白的表达水平以维持体内的铁平衡,而其本身的表达则受到负调控因子Fur(ferric uptake regulator)的调节.在体内,RyhB与Hfq蛋白和核糖核酸酶E (ribonuclease E, RNase E)形成核蛋白复合物sRNP来发挥活性.sRNP通过RyhB与靶基因的互补配对序列作用于靶基因的核糖体结合位点,阻断靶mRNA的翻译,并迅速引起靶mRNA的降解.此外,RyhB还可以通过影响致病菌的生物膜形成、趋化性、耐酸性等方面的能力对细菌的致病力进行调节.本文综述了RyhB的结构、功能及作用机制方面的研究进展,并对其存在的生理意义进行了探讨.     

非编码RNA和RNA沉默

生物体内存在大量的非编码RNA ,它们形态各异 ,功能也千差万别 ,在生物的生长、发育、分化进程中扮演着不同的角色 ,尤其是siRNA ,它是RNA沉默的诱因。RNA沉默是真核生物特有的现象 ,它需要一系列因子的参与 ,其中RNA依赖性的RNA聚合酶是沉默起始的关键 ,Dicer酶是形成siRNA的基础 ,而RNA沉默诱导复合体 (RSIC)等是发生RNA沉默“链式反应”的关键因子     

小RNA与蛋白质的相互作用

小分子调控RNA,包括siRNA（small interfering RNA）、miRNA（microRNA）和piRNA（piwiinteracting RNA）、hsRNA（heterochromatin associatedsmall RNA）等,是当前生命科学研究的前沿热点。越来越多的证据表明,这些小分子RNA存在于几乎所有较高等的真核生物细胞中,对生物体具有非常重要的调控功能。它们通过各种序列特异性的RNA基因沉默作用,包括RNA干扰（RNAi）、翻译抑制、异染色质形成等,调控诸如生长发育、应激反应、沉默转座子等各种各样的细胞进程。随着对这些小分子调控RNA的发现,一些RNascⅢ酶家族成员、Argonaute蛋白质家族成员及RNA结合蛋白质等先后被鉴定为小RNA的胞内蛋白质合作者,参与小RNA的加工成熟和在细胞内行使功能。本综述简介一些RNA沉默作用途径中重要组分的结构和功能的研究进展。