21世纪的RNA研究

美国<科学>周刊分别于2001年、2002年初评出上一年度的世界十大科学突破,RNA研究均位居第二:生命可能始于RNA,而非DNA;RNA干扰在使哺乳动物细胞的基因表达沉默和RNA在调控方面的很多新发现超出了科学家原先的想象.国际上,RNA研究正在受到人们前所未有的关注.     

AC2 and AC4 proteins of <Emphasis Type="Italic">Tomato yellow leaf curl China virus</Emphasis> and <Emphasis Type="Italic">Tobacco curly shoot virus</Emphasis> mediate suppression of RNA silencing

Tomato yellow leaf curl China virus Y10 isolate (TYLCCNV-Y10) alone could systemically infect host plants such as Nicotiana benthamiana without symptoms. In contrast, Tobacco curly shoot virus Y35 isolate (TbCSV-Y35) alone induces leaf curl symptoms in N. benthamiana. When inoculated into transgenic N. benthamiana plants expressing GFP gene (line 16c), TYLCCNV-Y10 neither reverses the established GFP silencing nor blocks the onset of GFP silencing. In contrast, TbCSV-Y35 can partially reverse the established GFP silencing and block the onset of GFP silencing in new leaves. In the patch co-infiltration assays, the AC2 and AC4 proteins of TYLCCNV-Y10 and TbCSV-Y35 could suppress local GFP silencing and delay systemic GFP silencing, suggesting that they are suppressors of RNA silencing. Comparison of the accumulation levels of GFP mRNA in the co-infiltration patches showed that Y10 AC2 and Y35 AC2 proteins had similar efficiency for suppression of RNA silencing. However, Y35 AC4 protein functioned as a stronger suppressor of RNA silencing than Y10 AC4 protein. There-fore, the pathogenicity difference between TbCSV-Y35 and TYLCCNV-Y10 may be related to the functional difference in their AC4 proteins.     

Lymphotactin enhances the in-vitro immune efficacy of dendritoma formed by dendritic cells and mouse hepatocellular carcinoma cells

Objective: To investigate the in-vitro antitumor immune responses of dendritoma formed by mouse hepatocellular carcinoma (HCC) cells and lymphotactin (Lptn) gene modified dendritic cells (DCs). Method: DCs prepared from mouse bone marrow were genetically modified by lymphotactin adenovirus, and fused with H22 cells by polyethylene glycol (PEG). RT-PCR and ELISA were employed to identify lymphotactin expression at mRNA and protein level. Cell phenotypes and fusion efficiency was detected by FACS. The stimulatory effect of DC on T cells was detected by mixed lymphocyte reaction. The cytotoxicity activity against H22 cells was assayed by LDH method. Results: Lymphotactin could be efficiently expressed by DCLptn/H22 hybridoma. DCLptn/H22 cells could induce potent T cell proliferation effect and generate strong cytotoxic T lymphocyte (CTL) reaction against allogenic H22 cells. Conclusion: Lymphotactin genetic modification could enhance the in vitro immune activity of the dendritoma.     

cDNA cloning and functional analysis of 4-coumarate --CoA: ligase (4CL) gene in Chinese white aspen

cDNA encoding 4-coumarate: CoA ligase (4CL) was isolated from the secondary developing xylem of Chinese white aspen (Populus tomentosa) by RT-PCR for the first time, which was 1619 bp in length. The coding sequence and putative amino acid sequence showed 97.53% and 97.00% identity to that of Pt4CL1 in quaking aspen (P. tremuloids), respectively. Molecular analysis indicated that 4CL was encoded by multiple genes in P.tomentosa, its mRNA was highly accumulated in xylem and the expression of 4CL revealed a biphasic pattern in one growing season, almost in phase with the expression of other related enzymes in lignin biosynthesis. Transgenic research showed that expression of antisense 4CL cDNA led to the decreasing of lignin content in transgenic tobaccos, among which the average reduction was 10.3% and the highest could be up to 18.9%. These data suggested that 4CL gene was a potential gene used in altering lignin biosynthesis by biotechnology for producing new materials of papermaking.     

RNA二级结构预测的模糊模型

基于模糊集合理论，提出了RNA二级结构预测的模糊模型．该模型通过状态空间的模糊分割以及模糊目标的引入等，可有效利用模糊动态规划算法给出该模糊模型的最优决策序列，并进而获得待预测RNA最优与次优的二级结构．基于模糊模型的方法具有许多优点，如计算复杂性的降低，最优与多个次优二级结构的一并获得，以及定性先验知识的有效融入等．完整地给出了RNA二级结构的模糊模型及其计算方法，并进行了具体的实现．将一个具体的BJK模糊模型结构实际应用于tRNA及tmRNA的数据集中，并与基于最小自由能的mfold工具以及基于SCFG的BJK文法模型进行了比较研究．实验结果表明该模型的有效性，相应的预测精度得到了进一步的提高．     

一个快速进化的microRNA家族

microRNAs(miRNAs)是长约22nt的非编码RNA,它可通过降解mRNA或抑制其翻译来调控基因。本文简要介绍了miRNAs的研究背景,阐述了通过比较近缘物种中非保守的miRNA基因来研究miRNAs分子进化的理由。我们的研究结果证实miRNA基因会受到达尔文正选择的影响,发生快速的进化。这一结果将为我们了解非蛋白编码基因的功能与进化提供崭新的视角。     

香蕉不同组织中总RNA提取方法的研究

针对香蕉组织中多糖、多酚等次生物质含量高的特点，以香蕉根、茎、叶、果实为试材，对其总RNA提取进行研究．比较了SDS法、改良SDS法、改良CTAB法和Trizol试剂盒法提取总RNA的效果，结果分析表明：改良SDS法能从香蕉根、茎、叶中获得质量高、完整性较好的总RNA，而改良CTAB法对果实的提取效果较佳，所得A260／A280均高于1．8，A260／A230大于2，0，28 S rRNA带的亮度约是18 S rRNA的2倍．其产率在根、茎、叶中均在400μg·g^-1以上，果实中可达49．34μg·g-^1.以上2种方法所得的总RNA均能满足RTPCR、Northern blot和cDNA文库建立等分子生物学研究，且具有快速、简便易行、成本较低的特点，适合于富含多糖、多酚的植物材料．     

小干涉RNA介导的RNA干涉机制及其应用的研究进展

RNAi系统是由SiRNA、沉默复合物、核酸酶、Argonaute蛋白家族的蛋白质组成,它具有高效性、特异性、持久性和遗传性等特征。RANi原理一般分为两个阶段:起始阶段和效应阶段,在siRNA存在的情况下,把目的mRNA降解成多个小片段RNA。RNAi在实际中可应用于基因功能的研究、基因治疗的研究、移植免疫耐受性的研究等,前景广阔。     

miRNA作用通路研究

后基因组时代的分子生物学研究需要从整体的角度,系统地阐述基因参与的生物调控过程,已有的通过构建基因调控网络的方式所做的系统生物学研究主要关注于蛋白编码基因之间的相互作用。随着近年来非编码RNA（noncoding RNA）,特别是miRNA研究的深入,显示生物体内存在着广泛的基因转录后调控。本论文通过建立综合蛋白质编码基因与miRNA基因相互作用关系的基因调控网络,分析了人类基因组中涉及miRNA的三类作用模式：（1）宿主基因与内含子miRNA共同作用于另一个蛋白编码基因。（2）miRNA簇中的不同miRNA分别作用于存在着相互作用的两个蛋白编码基因。（3）由两个宿主基因与其各自的内含子miRNA形成双向负调控回路。本研究的结果为进一步认识人类miRNA基因的功能特性提供了重要参考,研究所预测的数据为实验验证提供了依据。