基因功能研究的一些新思路

2001年人类基因组计划、美国塞莱拉遗传信息公司在美国《科学》杂志和英国《自然》杂志联合宣布,他们绘制出了准确、清晰、完整的人类基因组图谱。基因组计划完成后新的难题摆在了科学家的面前,就是基因功能的研究,以及怎样提高基因功能研究的效率。转基因技术较好地继承了生理学原有的研究思路,可能成为基因与整体功能联系起来的重要研究方法之一。成为今后大规模进行基因功能研究的有效手段。     

基因功能研究方法浅介

随着人类基因组计划的进展,数据库中积累了越来越多未知功能的基因序列,分析这些基因的功能将成为基因组计划的主要任务。本介绍了几种研究特定基因功能的方法及程序。     

植物病毒诱导的基因沉默在基因功能研究中的应用

传统的植物遗传转化方法周期长、工作量大、过程繁琐,不利于基因功能的快速高通量鉴定．近年来随着基因沉默机制研究的深入和不断发展,利用病毒诱导的基因沉默(Virus induced gene silencing,VIGS)进行植物功能基因组研究作为一种快速、高通量的反向遗传学工具已被广泛应用在烟草、马铃薯、番茄等植物中, 在大规模的植物基因组功能鉴定中展示了广阔的应用前景．综述了 VIGS 的作用机制、植物病毒栽体、转化方法以及在植物基因功能研究等方面的应用及前景．     

病毒诱导的基因沉默及其在植物功能基因组研究中的应用

病毒诱导的基因沉默已成为研究植物功能基因组的重要工具. VIGS 体系因其方法简便、周期性短以及避免植物转化等诸多优点, 已在利用正向遗传学和反向遗传学寻找和鉴定基因功能方面发挥了日益重要的作用. 越来越多的植物病毒被改造成为VIGS 载体, 并已在植物发育、生物逆境、非生物逆境、细胞代谢、信号传导等基因功能研究方面得到了应用. 本文围绕VIGS的发展以及在植物功能基因鉴定中的应用及前景提出了展望.     

RNA干扰文库及其在功能基因组学研究中的应用

随着人类基因组大规模测序的完成,下一步的挑战是了解每一个基因的功能 . RNA 干扰文库为大规模基因功能筛选提供了可能 . 虽然用于线虫等模式生物的 RNAi 文库,已经证明是大规模基因功能筛选的有效方法,但这些文库不能用于高等动物的细胞 . 自 2003 年以来,用于人的细胞和哺乳动物细胞的 RNAi 文库取得了突破,相继出现构建已知基因 RNAi 文库和构建随机 RNAi 文库的报道,并成功地应用于大规模基因功能的筛选 . RNAi 文库作为一种简单、高效、大规模、高通量的功能基因组学研究的工具,将在基因功能研究、发现新的药物靶基因、发现疾病相关基因等方面有广阔的应用前景 .     

酵母基因中断技术

酵母基因中断技术是研究酵母基因功能的重要手段,自80年代初诞生以来经历了不断的改进和发展.PCR介导的酵母基因中断技术,大大简化了操作,实现了酵母基因的精确缺失;酵母基因的多重中断技术,可在酵母内实现多个基因的中断;可进行大规模基因中断和功能分析的酵母基因中断技术,适应了在酵母全基因组测序完成的情况下进行功能基因组学研究的要求.酵母基因中断技术对人类基因功能研究也有很大启示作用.     

基因功能研究的技术和方法

随着基因组计划的深入,越来越多的有生理意义的基因被成功克隆,对基因功能的研究显得日益重要。目前基因功能研究的主要方法有;基因转导,反义技术,核酶,基因重组,染色体转导技术等。     

酵母基因中断技术

酵母基因中断技术是研究酵母基因功能的重要手段，自80年代初诞生以来经历了不断的改进和发展.PCR介导的酵母基因中断技术，大大简化了操作，实现了酵母基因的精确缺失；酵母基因的多重中断技术，可在酵母内实现多个基因的中断；可进行大规模基因中断和功能分析的酵母基因中断技术，适应了在酵母全基因组测序完成的情况下进行功能基因组学研究的要求.酵母基因中断技术对人类基因功能研究也有很大启示作用.     

微生物全基因组测序研究进展

本文综述了近年来大规模微生物基因组核酸序列测定的最新研究进展,介绍微生物全基因组测序的基本方法、序列的收集组装,序列缺口的填补,以及序列资料的计算机分析整理。大规模基因组测序完成后,未来面临的更大挑战是在ＤＮＡ序列基础上认识微生物的完整生物学功能。为此本文也介绍了有关基因功能分析的新技术,并对微生物基因组功能分析的未来发展作了展望。     

基因组水平的基因挖掘与功能分析方法的探讨

为了实现基因组中特定基因功能的注释,研究者提出一种新的思路,即利用对目的基因启动子上游的顺式元件的功能的分析,进一步来推断目的基因的功能。在此主要对基因组水平的基因挖掘与功能分析方法及其研究进展进行了探讨。     

面向大规模人群的基因组注释系统

基因组注释是识别出基因组序列中功能组件的过程,其可以直接对序列赋予生物学意义,由此方便研究者探究和分析基因组功能.基因组注释可以帮助研究从三个层次上理解基因组,一种是在核苷酸水平的注释,主要确定DNA序列中基因、RNA、重复序列等组件的物理位置,包括转录起始,翻译起始,外显子边界等具体位置信息.同时可以注释得到变异在不...     

Using Targeted Large Deletions and High-EfficiencyN-Ethyl-N-nitrosourea Mutagenesis for Functional Analyses of the Mammalian Genome

The Human Genome Project has generated nucleotide sequences from an estimated 80,000 to 100,000 genes, only a small fraction of which have a known role. Nucleotide sequence information alone is insufficient to predict gene function. One of the most powerful ways of revealing gene function, as demonstrated in bacteria, worms, yeast, and flies, is to generate mutations and characterize them at both the phenotypic and the molecular levels. Given the physiological and anatomical parallels between mouse and human, genotype–phenotype relationships established in mice can be extrapolated to human syndromes. A new method is described for functional genetic analyses in the mouse that uses loxP/Cre engineering to generate coat color-tagged large deletions. The haploid regions can then be dissected by mutagenesis withN-ethyl-N-nitrosourea in phenotype-driven screens to obtain functional information on genes in any desired region of the mouse genome.     

Generation of Radiation-Induced Deletion Complexes in the Mouse Genome Using Embryonic Stem Cells

As the genetic and physical mapping stage of the Human Genome Project nears completion, the focus is shifting toward the development of technologies for high-throughput analysis of gene function. Whereas DNA sequencing will enable the assignment of presumed function to a large number of genes in mice and humans, it is clear that the great majority of genes will have to be evaluatedin vivoto accurately assess their role in a complex organism. While gene targeting in mouse embryonic stem (ES) cells is the current method of choice for the characterization of gene function in mice, it remains relatively labor intensive and lacks the throughput required for analysis of genome function on a large scale. Alternative methods of efficient mutagenesis will clearly be required for this task. Chromosomal deletions are powerful tools in the genetic analysis of complex genomes, enabling the systematic identification and localization of functional units along defined chromosomal regions. Not only are deletions useful for the identification of genetic functions, but they serve as mapping reagents for existing mutations or traits. While their use has been an essential tool inDrosophilagenetics, classical mutagenesis in mice has been logistically impractical for generating deletions. We have previously described an efficient method for generating radiation-induced deletion complexes at defined regions in the genome using ES cells. In this article, we detail the methodological aspects of this technology and describe the applications of chromosomal deletions for characterizing gene function in ways that make optimal use of the information generated by the first stage of the Genome Project.     

一种高效的功能基因分离解析法

人类基因组计划（Human Genome Project）的实施揭开了各种生物基因组解析的序幕［1~3］。随着各种生物的基因组解析的顺利进行,遗传基因的功能研究以及寻找新的功能基因变得越来越重要。本文介绍的MegacloneTM技术、MegasortTM技术［4］以及MPSS技术［5］可以高效地分离解析各种功能基因。 Abstract:The implementation of the Human Genome Project preludes the analyzing of biologic genomes［1~3］.Following the successful analysis of diverse biologic genomes,it becomes more and more important to research the functions of genes and to find new functional genes.In this article,we use the techniques of MegacloneTM,MegasortTM［4］ and MPSS［5］ to sort and sequence effectively different functional genes.     

C. elegans neuroscience: genetics to genome

From their earliest experiments, researchers using Caenorhabditis elegans have been interested in the role of genes in the development and function of the nervous system. As the C. elegans Genome Project completes the genomic sequence, we review the accomplishments of these researchers and the impact that the Genome Project has had on their research. We also speculate on future directions in this research that are enabled by the efforts of the Genome Project.     

Pichia stipitis genomics, transcriptomics, and gene clusters

Genome sequencing and subsequent global gene expression studies have advanced our understanding of the lignocellulose-fermenting yeast Pichia stipitis . These studies have provided an insight into its central carbon metabolism, and analysis of its genome has revealed numerous functional gene clusters and tandem repeats. Specialized physiological traits are often the result of several gene products acting together. When coinheritance is necessary for the overall physiological function, recombination and selection favor colocation of these genes in a cluster. These are particularly evident in strongly conserved and idiomatic traits. In some cases, the functional clusters consist of multiple gene families. Phylogenetic analyses of the members in each family show that once formed, functional clusters undergo duplication and differentiation. Genome-wide expression analysis reveals that regulatory patterns of clusters are similar after they have duplicated and that the expression profiles evolve along with functional differentiation of the clusters. Orthologous gene families appear to arise through tandem gene duplication, followed by differentiation in the regulatory and coding regions of the gene. Genome-wide expression analysis combined with cross-species comparisons of functional gene clusters should reveal many more aspects of eukaryotic physiology.     

Combining functional genomics and chemical biology to identify targets of bioactive compounds

Genome sequencing projects have revealed thousands of suspected genes, challenging researchers to develop efficient large-scale functional analysis methodologies. Determining the function of a gene product generally requires a means to alter its function. Genetically tractable model organisms have been widely exploited for the isolation and characterization of activating and inactivating mutations in genes encoding proteins of interest. Chemical genetics represents a complementary approach involving the use of small molecules capable of either inactivating or activating their targets. Saccharomyces cerevisiae has been an important test bed for the development and application of chemical genomic assays aimed at identifying targets and modes of action of known and uncharacterized compounds. Here we review yeast chemical genomic assays strategies for drug target identification.