植物功能基因组学及其研究技术

植物基因组的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.植物功能基因组学研究是利用结构基因组学积累的数据,从中得到有价值的信息,阐述DNA序列的功能,从而对所有基因如何行使其职能并控制各种生命现象的问题作出回答.近年来植物功能基因组学的研究技术主要包括表达序列标签、基因表达的系列分析、DNA微阵列和反向遗传学等.对植物功能基因组学的研究将有利于我们对基因功能的理解和对植物形状的定性改造和利用.     

植物功能基因组学研究

基因组研究已进入了以功能基因组研究为特征的后基因组时代.本文介绍了基因组及基因组学的概念及功能基因组学的研究方法,并综述了植物功能基因组学的研究进展.     

卵菌基因组学研究进展

卵菌是一类真核生物,包括许多植物病原菌,引起许多作物、花卉等发生灾难性病害.卵菌具有独特分类地位的群体,由于表现出丝状等特性传统上被划分到真菌界中.近年来,随着基因组学理论和技术的发展,大量的卵菌基因组相关的数据库被建立.本文综述了现有的卵菌基因组资源以及卵菌分子遗传学的研究进展.重点论述了卵菌基因组的大小、组成和不稳定性以及卵菌的基因结构与功能基因组等五个方面,并分析了卵菌基因组的特点.     

MoFLP1, encoding a novel fungal fasciclin-like protein, is involved in conidiation and pathogenicity in Magnaporthe oryzae

Fasciclin family proteins have been identified as cell adhesion molecules in various organisms. In this study, a novel Magnaporthe oryzae fasciclin-like protein encoding gene, named MoFLP1, was isolated from a subtractive suppressive cDNA library and functionally analyzed. Sequence analysis showed that the MoFLP1 gene contains an open reading frame (ORF) of 1050 nucleotides encoding 349 amino acids with a calculated molecular weight of 35.85 kDa and a pl of 7.76. The deduced MoFLP1 protein contains a 17-amino acid secretion signal sequence and an 18-amino acid sequence with the characteristics of a glycosylphosphotidylinositol (GPI) anchor additional signal at its N- and C-terminuses, respectively. Potential N-glycosylation sites and domains involving cell adhesion were also identified in MoFLP1. Sequence analysis and subcellular localization by the expression of MoFLP1-GFP fusion construct in M. oryzae indicated that the MoFLP1 protein is probably localized on the vacuole membrane. Two MoFLP1 null mutants generated by targeted gene disruption exhibited marked reduction ofconidiation, conidiai adhesion, appressorium turgot, and pathogenicity. Our results indicate that fasciclin proteins play important roles in fungal development and pathogenicity in M. oryzae.     

Agrobacterium tumefaciens-mediated genetic transformation of the phytopathogenic fungus Penicillium digitatum

Agrobacterium tumefaciens-mediated transformation (ATMT) system was assessed for conducting insertional mutagenesis in Penicillium digitatum, a major fungal pathogen infecting post-harvest citrus fruits. A transformation efficiency of up to 60 transformants per 106 conidia was achieved by this system. The integration of the hph gene into the fungal genome was verified by polymerase chain reaction (PCR) amplification and sequencing. These transformants tested were also shown to be mitotically stable. Southern blot analysis of 14 randomly selected transformants showed that the hph gene was randomly integrated as single copy into the fungal genome of P. digitatum. Thus, we conclude that ATMT of P. digitatum could be used as an alternatively practical genetic tool for conducting insertional mutagenesis in P. digitatum to study functional genomics.     

Investigation of the biological roles of autophagy in appressorium morphogenesis in Magnaporthe oryzae

Magnaporthe oryzae has been used as a primary model organism for investigating fungus-plant interaction. Many researches focused on molecular mechanisms of appressorium formation to restrain this fungal pathogen. Autophagy is a very high conserved process in eukaryotic cells. Recently, autophagy has been considered as a key process in development and differentiation in M. oryzae. In this report, we present and discuss the current state of our knowledge on gene expression in appressorium formation and the progress in autophagy of rice blast fungi.     

七种植物病原真菌对人参皂苷Rb1生物转化的初步研究

利用7种植物病原真菌对人参皂苷Rb1进行生物转化,通过薄层层析法检测,结果表明：有6种真菌能将人参皂苷Rb1转化为Rd1但只有齐整小核菌的转化高效而彻底,其他几种转化效率低或副产物多。     

水稻T-DNA插入突变技术研究

随着水稻基因组全序列的测定完成,功能基因组学已成为重点研究内容.农杆菌介导的T-DNA标签法是近年发展起来的一种有效的分子生物学技术.它具有程序简便,转化效率高,大规模转化等特点.阐述了T-DNA标签法及其改进后的激活标签技术和增强子技术的特点,并介绍了T-DNA插入突变在水稻突变体库的建立中所取得的研究进展,从而表明T-DNA插入突变技术是水稻功能基因组研究的一个有效途径.     

聚合酶链反应检测真菌感染的实验研究

本研究以NS5-NS6为引物,扩增真菌同源性序列ssu-rDNA片段,建立广范围真菌检测的方法。用此方法扩增医学主要致病真菌、细菌和人体细胞,结果所有真菌均扩增出一个约310bp的产物,而细菌和人体细胞均扩增阴性,1pg白色假丝酵母菌DNA即可检出。     

植物病原真菌侵染过程动画模拟的教学实践

植物病理学是研究微观和宏观世界客观规律的学科,是针对由真菌、细菌、病毒、植物寄生线虫等生物因素引起的侵染性病害而言的,其研究对象主要是人们肉眼看不见的病原微生物及寄主植物受害后的病理现象。结合本学科的最新研究成果,采用动画模拟植物病原物侵染植物的微观过程来展示知识点,化难为易,可培养学生的学习兴趣和积极性,提高了教学效果,准确地达到了预定的教学目标。介绍了植物病原菌侵染过程动画制作的基本过程,分析了近10年来植物病原菌侵染过程动画模拟方法的教学效果,总结了相关经验,并对植物病原菌侵染过程动画模拟教学方法中存在的一些问题进行了讨论。     

A simple and effective method for total RNA isolation of appressoria in Magnaporthe oryzae

Appressorium formation is an important event in establishing a successful interaction between the rice blast fungus, Magnaporthe oryzae, and its host plant, rice. An understanding of molecular events occurring in appressorium differentiation will give new strategies to control rice blast. A quick and reliable method to extract total RNA from appressorium is essential for studying gene expression during appressorium formation and its mechanism. We found that duplicate film is an efficient substratum for appressorium formation, even when inoculated with high density conidia. When inoculated with conidia at 1×106ml-1, the percentages of conidium germination and appressorium formation were (97.98±0.67)% and (97.88±0.45)%, respectively. We applied Trizol before appressorium collection for total RNA isolation, and as much as 113.6 pg total RNA was isolated from the mature appressoria at 24 h after inoculation. Functional analysis of two genes, MNH6 and MgATG1, isolated from the cDNA subtractive library, revealed that the quantity of RNA was good enough to construct a cDNA (complementary DNA) library or a cDNA subtractive library. This method may be also applicable for the appressorium RNA isolation of other pathogenic fungi in which conidia differentiate into appressoria in the early stages of host infection.     

Application of cDNA array for studying the gene expression profile of mature appressoria of Magnaporthe grisea

Appressorium is an infection structure of the phytopathogenic fungus Magnaporthe grisea. Analysis of gene expression profiles ofappressorium development provides insight into the molecular basis of pathogenicity and control of this fungal plant disease. A cDNA array representing 2927 unique genes based on a large EST （expressed sequence tag） database ofM. grisea strain Y34 was constructed and used to profile the gene expression patterns at mycelium and appressorium maturation stages. Compared with mycelia, 55 up-regulated and 22 down-regulated genes were identified in mature appressoria. Among 77 genes, 16 genes showed no similarity to the genome sequences of M. grisea. A novel homologue of peptidyl-prolyl cis-trans isomerase was found to be expressed at low-level in mature appressoria of M. grisea. The results indicated that the genes such as pyruvate carboxylase, phospholipid metabolism-related protein and glyceraldehyde 3-phosphate dehydrogenase involved in gluconeogenesis, lipid metabolism and glycolysis, showed differential expression in mature appressoria. Furthermore, genes such as PTHll, beta subunit of G protein and SGTI involved in cell signalling, were expressed differentially in mature appressoria. Northern blot analysis was used to confirm the cDNA array results.     

1,3,4-噁二唑类衍生物的合成及杀菌活性研究

杀菌剂是现代农药的一个重要组成部分,利用它,人们可以把作物的生长控制在对人有利的方向。为了寻找并筛选具有较高杀菌活性的化合物,将具有优良生物活性的1,3,4-噁二唑结构引入化合物中,合成20个目标化合物。本文采用离体活性测试法对目标化合物进行杀菌活性测试,获得对部分植物病原真菌具有较高活性和应用价值的杀菌剂。     

茴香醛抗真菌作用的实验研究

本文以琼脂稀释法测定了茴香醛对10种真菌的最低抑菌浓度.实验发现茴香醛对所试10种真菌均有抑和杀菌作用,对其中自色念珠菌、新型隐球菌及孢子丝菌等条件致病真菌也具有抗真菌活性.其临床意义有待于进一步探讨。     

转座子及其在水稻基因组学中的研究进展

本文介绍了转座子的基本结构和转座子标签技术分离基因的基本原理、方法;并就转座子标签技术的研究新进展,转座子标签技术在水稻功能基因组学研究中的应用等方面进行了综述.     

A botanist,mycorrhiza and a knighthood

Most plants are attached via their roots to specialized fungi. The filamentous hyphae of these fungi extend beyond the nutrient-depleted zone in the soil, absorbing and supplying nutrients to the plants in exchange for photosynthetically-made carbon compounds manufactured by the plant. The hyphae also attach to litter and decompose the organic matter, releasing the mineral ions sequestered in the structural polymers. The mineral ions are absorbed and translocated to the roots. Most plants are so dependent on their fungal partners for supplying nutrients that they languish or die without them. Our understanding of this fascinating interaction, to a large extent, has come from the pioneering work of Sir David J Read, FRS, Emeritus Professor of Plant Sciences at the University of Sheffield, UK. Ramesh Maheshwari is a former Professor and Chairman of the Department of Biochemistry, Indian Institute of Science, Banglaore. This is his sixth article for Resonance.     

短毛独活精油提取及抑菌作用研究

用石油醚提取获得短毛独活精油,采用生长速率法测试短毛独活精油的杀菌活性。结果表明:短毛独活精油对12种供试真菌均具有不同程度的抑菌作用,其中对小麦纹枯病菌的抑制率最强,达到100%;其次是油菜菌核病菌、辣椒疫霉病菌,达到80%以上;而对玉米大斑病菌、稻瘟病菌的抑制率相对较低。     

植物几丁质酶及其基因工程研究进展

几丁质是构成大多数真菌细胞壁的主要成分.几丁质酶广泛存在于植物、动物及微生物细胞和组织中,参与多种生理过程.研究发现,许多微生物都可以产生几丁质酶,几丁质酶的生物活性可显著抵抗植物真菌病害.目前有多种植物、微生物几丁质酶基因已经被克隆出来,并有部分已经转化应用于植物抗病,有些显现出良好效果.     

Comparative genomics

An important hallmark of biological research is the aspect of ‘comparisons’. As the complete genome sequences of numerous organisms have become available, the emphasis in biology has shifted to comparisons at the genome level. Indeed, the last few years have witnessed an exponential rise in the number of organisms whose complete genome has been sequenced, and we are still climbing up the graph. The present article, a primer, explains how one can extract a great deal of information from such analyses that is of great value in our research. The subject of comparative genomics impinges on evolutionary biology and phylogenetic reconstructions of the tree of life, drug discovery programs, function predictions of hypothetical proteins and genes, regulatory motifs and other non-coding DNA motifs, and genome flux and dynamics. Finally the article describes how the information one can extract from a comparative analysis of genomes depends to a large extent, on the specific aspect of the genomes that is being compared and the phylogenetic distances of the organisms involved.