抑制消减杂交法分离紫花苜蓿幼苗铝胁迫诱导表达的cDNA

利用抑制消减杂交(SSH)技术分离铝胁迫诱导紫花苜蓿差异表达的基因,以水培试验获取的中苜一号幼苗为材料,以80μmol/L铝离子胁迫的紫花苜蓿作为试验组,未胁迫的为驱动组,构建了一个包含456个克隆的SSH文库。对构建的文库进行鉴定,随机选取20个阳性克隆测序,共获得15条有效EST序列,然后将测序结果提交到GenBank进行Blastn比对,获得了3条未知基因的序列,推测它们可能与植物的抗铝作用有关。检测结果表明,构建的文库质量较好,可以进行进一步深入研究,为揭示植物耐铝性的分子机理提供理论基础。     

植物抗旱和耐重金属基因工程研究进展

干旱和重金属污染严重影响植物的生长发育.植物耐逆相关基因的克隆和功能鉴定研究,为通过基因工程途径提高植物的抗逆性奠定了理论基础.水分亏缺、高盐、低温和重金属胁迫都能诱导LEA(late embryogenesis abundant protein)基因的表达.转基因研究表明,LEA蛋白具有抗旱保护作用、离子结合特性以及抗氧化活性;水孔蛋白存在于细胞膜和液泡膜上,在细胞乃至整个植物体水分吸收和运输过程中发挥重要作用.干旱和盐胁迫促进水孔蛋白基因转录物的积累.过量表达水孔蛋白可增强水分吸收和运输,提高植物的抗旱能力.金属转运蛋白参与重金属离子的吸收、运输和累积等过程.这些蛋白基因在改良草坪草植物的抗旱节水和耐重金属能力等方面具有潜在的应用价值.     

植物响应盐胁迫组学研究进展

盐胁迫对植物生长的影响主要表现在离子毒害、渗透胁迫以及次级氧化胁迫等,植物遭受盐胁迫时迅速启动相关基因,进行转录调控,进而合成相应蛋白质来控制代谢物合成和离子转运以调节渗透平衡。随着现代分子生物学迅速发展,对植物耐盐机理研究也深入到了转录组、蛋白质组、代谢组及离子组等水平,"组学"研究为耐盐基因鉴定及标志性代谢物的挖掘等提供了有力手段。该文对近年来国内外有关转录组学、蛋白质组学、代谢组学、离子组学的主要研究方法及在盐胁迫中的应用研究进展进行综述,以揭示植物耐盐机理,为优良耐盐碱植物的筛选与培育提供支持。     

丹波黑大豆GmbHLH30转录因子耐铝功能初步研究

铝毒是酸性土壤作物生长的主要限制因素。前期研究发现,铝胁迫下,耐铝型丹波黑大豆SSH(suppression subtractive hybridization,SSH)cDNA文库中bHLH30转录因子基因上调表达,推测该基因与丹波黑大豆耐铝性相关。克隆GmbHLH30基因,构建GmbHLH30植物表达载体pK2-35S-GmbHLH30,并在烟草中过量表达获得转GmbHLH30的转基因烟草植株。在铝胁迫下,转GmbHLH30的转基因烟草相对根伸长率比野生型烟草大,可溶性糖和脯氨酸含量高,H2O2水平低。表明GmbHLH30基因的过量表达可以增强植物的耐铝能力,暗示GmbHLH30转录因子参与调控植物的耐铝特性。     

植物适应铝毒胁迫的生理及分子生物学机理

铝毒是酸性土壤上限制作物生长最重要的因素,严重影响着全世界和中国大约40％和21％耕作土壤的作物生产．近几十年来,世界各国针对植物的铝毒及其耐铝机制进行了大量的研究,并取得了较大进展．文中重点综述了植物适应铝胁迫基因型差异筛选方法及其鉴定技术、植物适应铝胁迫的生理基础及分子生物学机制等方面的研究进展,简要讨论了今后的研究方向．     

干旱胁迫诱导下植物基因的表达与调控

干旱胁迫能够诱导植物表达大量的基因 ,研究这些基因的表达与调控 ,为植物抗旱的定向育种创造条件。本文系统介绍了在干旱胁迫条件下 ,植物体内渗透调节物质和可溶性糖合成有关的基因、离子和水分通道及Lea蛋白基因的表达 ,以及与这些基因表达相关的调控元件和因子 ,干旱胁迫信号转导等方面的最新研究进展。     

植物铝胁迫响应基因的研究进展

铝毒是酸性土壤中植物生长和作物生产的主要限制因子.近年来的很多研究应用差异显示PCR、抑制差减cDNA文库和DNA微正列等技术,在一些铝耐受型和敏感型植物中鉴定了很多铝胁迫响应基因.本研究通过参阅国内外有关报道和结合本实验室的研究成果,从铝诱导的通道蛋白、代谢相关、胁迫和细胞死亡以及信号转导相关基因4个方面的研究进展进行了综述.     

用基因芯片技术分析铝胁迫下小麦的基因表达谱

目的:采用基因表达谱分析方法,探讨小麦耐铝的分子机理。方法:利用抑制消减杂交(SSH)技术,以小麦的铝敏感品种Chisholm及其耐铝近等基因系Chisholm-T(其耐铝性来自小麦品种Atlas66)的根尖为材料,构建了2个铝胁迫后的SSHcDNA文库,共含有1628个表达序列标签(EST),利用这些EST制作了小麦根系的cDNA基因芯片。以cDNA基因芯片为平台,在铝胁迫后6h、1d、3d和7d,分别比较Chisholm和Chisholm-T之间的基因表达谱差异。结果:在各个时间点,耐铝和不耐铝小麦材料之间约有5%的EST表现出差异表达。对所有差异表达的EST进行测序分析,序列数据经Pipe-Online2.0进行毗连序列群(contig)拼接,发现只有8.3%的重复序列。结论:SSH是一种非常有效的差减和均一化的建库方法。对有功能注释的差异表达基因进行功能分类分析,表明这些基因参与了植物体内的电子传递、信号传导、植物保护和次生物质的代谢活动。     

cDNA-AFLP技术在植物耐盐基因鉴定上的应用

cDNA-AFLP技术是近年来广泛应用于植物基因分离与表达研究的mRNA指纹图谱技术,在分离植物特异性基因等生物技术研究中发挥了巨大作用.在植物耐盐基因鉴定方面,该技术的应用处于新兴初始阶段,但已取得了诸多成效.本文在概述植物耐盐响应机制的基础上,着重对利用cDNA-AFLP技术鉴定的植物耐盐相关基因及其相应作用机制进行了阐述,并对其在植物耐盐分子生物学研究上的应用前景进行了展望.     

植物抗盐胁迫研究进展

盐胁迫严重制约了农业生产,解析盐胁迫机理受到关注。随着抑制消减杂交和基因表达序列分析等大规模表达基因鉴定技术在抗盐研究中的应用,盐碱胁迫下功能基因的获得量迅速增加。综述了近年来在酵母、拟南芥、水稻等生物上利用基因芯片等方法进行抗盐机理研究取得的成果,并介绍了结合系统生物学的方法进行抗盐研究取得的进展,以及植物抗盐胁迫研究的发展前景。     

cDNA-AFLP技术及其在植物基因表达研究中的应用

cDNA-AFLP技术是一种新的研究基因表达的技术,具有重复性好、稳定、可靠的特点,可对生物体转录组进行全面、系统的分析,广泛应用于基因表达特性研究、植物遗传标记分析和分离植物基因等方面.近年来随着技术的不断进步,设备的不断改进,许多新的研究方法不断的产生,该技术取得了迅速的发展.本文就cDNA-AFLP技术的原理、技术特点及其在植物基因表达研究中的应用进行了综述.     

植物功能基因组学研究进展

植物基因组研究已经由以全基因组测序为目标的结构基因组学转向以基因功能鉴定为目标的功能基因组学研究.本简要介绍了植物功能基因组的主要研究方法,如基因表达系列分析法、表达序列标签法、差异表达谱基因芯片法、蛋白质组学分析法以及生物信息学等及其研究现状,并展望了植物功能基因组学的应用前景.     

Technical variables in high-throughput miRNA expression profiling: much work remains to be done

MicroRNA (miRNA) gene expression profiling has provided important insights into plant and animal biology. However, there has not been ample published work about pitfalls associated with technical parameters in miRNA gene expression profiling. One source of pertinent information about technical variables in gene expression profiling is the separate and more well-established literature regarding mRNA expression profiling. However, many aspects of miRNA biochemistry are unique. For example, the cellular processing and compartmentation of miRNAs, the differential stability of specific miRNAs, and aspects of global miRNA expression regulation require specific consideration. Additional possible sources of systematic bias in miRNA expression studies include the differential impact of pre-analytical variables, substrate specificity of nucleic acid processing enzymes used in labeling and amplification, and issues regarding new miRNA discovery and annotation. We conclude that greater focus on technical parameters is required to bolster the validity, reliability, and cultural credibility of miRNA gene expression profiling studies.     

植物次生代谢基因工程研究进展

随着对植物代谢网络日渐全面的认识,应用基因工程技术对植物次生代谢途径进行遗传改良已取得了可喜的进展.对次生代谢途径进行基因修饰的策略包括:导入单个、多个靶基因或一个完整的代谢途径,使宿主植物合成新的目标物质;通过反义RNA和RNA干涉等技术降低靶基因的表达水平,从而抑制竞争性代谢途径,改变代谢流和增加目标物质的含量;对控制多个生物合成基因的转录因子进行修饰,更有效地调控植物次生代谢以提高特定化合物的积累.作者结合对大豆种子异黄酮类代谢调控和基因工程改良的研究,着重介绍了花青素和黄酮类物质、生物碱、萜类化合物和安息香酸衍生物等次生代谢产物生物合成的基因工程研究进展.     

Gene amplification and expression by RNA viruses and potential for further application to plant gene transfer

The great majority of plant viruses encapsidate messenger-sense ssRNA and have no natural DNA phase in their life cycle. Despite their RNA nature, essentially any desired change can be introduced into such genomes by using recombinant DNA techniques with suitably constructed, expressible viral cDNA clones. For some viruses such as brome mosaic virus, these methods have been used to define the sequences controlling RNA-directed genomic RNA replication and the expression of internal genes via subgenomic mRNAs. The results suggest a surprising degree of genetic flexibility, which appears to be reflected in the varied gene complements and genetic organizations of presumably related plant and animal RNA viruses sharing conserved replication genes. Foreign genes inserted in such RNA virus genomes can be amplified and expressed to a high level in transfected plant cells. In addition to the potential use of such viruses as episomal expression vectors, it should be possible to couple the viral pathways of RNA-dependent RNA synthesis to amplify and to further regulate the expression of genes transformed into plant chromosomes.     

植物酸性转化酶基因及其表达调控

酸性转化酶是蔗糖代谢的关键酶,在植物体中具有重要的生理作用.近十几年来,许多植物酸性转化酶基因已经克隆,其基因表达调控的研究也取得了很大的进展.本文综述了植物酸性转化酶基因及其蛋白结构、基因表达的器官和发育特异性以及糖、受伤、病原、胁迫和激素对基因表达的调节和蛋白抑制因子对酶活性的影响,并讨论了当前在该研究领域存在的问题.     

Gene expression during the male gametophytic phase

Abstract

In recent years a number of experimental findings have indicated that in higher plants the gametophytic phase is able to express its own genetic information, a large part of which it shares with the sporophytic generation. Quantitative estimates of haploid and haplodiploid gene expression have been obtained by mRNA and isozyme analysis in several plant species: 60-70% of the genes are expressed in both pollen and plant, about 10% are pollen-specific, and 20% represent the sporophytic domain. Moreover, it has been demonstrated that stage-specific genes are expressed in the gametophytic generation: at least two sets of genes are activated during pollen development, others are expressed only in the postshedding period, during germination and tube growth. Studies have been made to ascertain the role played by gametophyte-expressed genes in pollen development; the in vivo and in vitro pollen tube growth rate has been revealed to be controlled by the gametophyte genome itself. Differential effects of specific chromosomal deficiencies on the development of maize pollen grains have indicated that components of normal microspore development are controlled by genes located in specific parts of the genome. For single gene analysis, gene transfer can be used; on the contrary, for traits with a multifactorial genetic control, direct proof of gene expression both in the gametophytic and the sporophytic generation can be obtained when selection is applied to the pollen population of a hybrid plant, and response to selection is observed in the resulting sporophytic progeny. Response to selection, applied at different stages of the gametophytic phase, has been described in the sporophytic progeny and this with regard to many adaptive traits; thus the phenomenon can have an important bearing on the genetic structure of natural populations and on higher plant evolution, it can also be used as a breeding tool to increase the efficiency of conventional selection methods.     

植物酸性转化酶基因及其表达调控

酸性转化酶是蔗糖代谢的关键酶,在植物体中具有重要的生理作用。近十几年来,许多植物酸性转化酶基因已经克隆,其基因表达调控的研究也取得了很大的进展。本文综述了植物酸性转化酶﹑ 基因及其蛋白结构、基因表达的器官和发育特异性以及糖、受伤、病原胁迫和激素对基因表达的调节和蛋白抑制因子对酶活性的影响,并讨论了当前在该研究领域存在的问题。     

Genome-wide co-expression based prediction of differential expressions

MOTIVATION: Microarrays have been widely used for medical studies to detect novel disease-related genes. They enable us to study differential gene expressions at a genomic level. They also provide us with informative genome-wide co-expressions. Although many statistical methods have been proposed for identifying differentially expressed genes, genome-wide co-expressions have not been well considered for this issue. Incorporating genome-wide co-expression information in the differential expression analysis may improve the detection of disease-related genes. RESULTS: In this study, we proposed a statistical method for predicting differential expressions through the local regression between differential expression and co-expression measures. The smoother span parameter was determined by optimizing the rank correlation between the observed and predicted differential expression measures. A mixture normal quantile-based method was used to transform data. We used the gene-specific permutation procedure to evaluate the significance of a prediction. Two published microarray data sets were analyzed for applications. For the data set collected for a prostate cancer study, the proposed method identified many genes with weak differential expressions. Several of these genes have been shown in literature to be associated with the disease. For the data set collected for a type 2 diabetes study, no significant genes could be identified by the traditional methods. However, the proposed method identified many genes with significantly low false discovery rates. AVAILABILITY: The R codes are freely available at http://home.gwu.edu/~ylai/research/CoDiff, where the gene lists ranked by our method are also provided as the Supplementary Material.     

The use of expression profiling to study pigment cell biology and dysfunction

Regulation of gene expression is a fundamental process by which cells respond to both intracellular and extracellular signals. For a pigment cell, alterations in gene expression regulate the processes of cell migration, lineage restriction, differentiation, type of pigment produced, and progression from a normal pigment cell to that of melanoma. To date, the identification of genes involved in normal pigment cell development has been accomplished by the cloning of individual mutant alleles, a single gene at a time. Current advances in technology have now made it possible to use expression profile analysis to investigate, on a genomic scale, the process of pigment cell development and function. This review compares and contrasts the methods of subtractive suppressive polymerase chain reaction (PCR) and differential display with that of cDNA microarray analysis.