华癸中慢生根瘤菌多铜氧化酶基因mco的功能研究

通过基因同源重组技术,构建华癸中慢生根瘤菌mco基因突变株HKmco,并探究了mco基因突变对根瘤菌生长、抗氧化以及共生固氮的影响.结果表明:mco基因缺失不影响华癸中慢生根瘤菌的生长能力,突变株对无机过氧化物H_2O_2不敏感,但对有机氧化物氢过氧化枯烯(CUOOH)非常敏感.植物盆栽实验表明,突变株HKmco感染紫云英宿主能形成红色有效根瘤,但mco基因突变株Hkmco的根瘤中的类菌体形态发生了改变,固氮酶活降低了29.8%.表明根瘤菌mco基因在抗氧化和共生固氮方面发挥了重要作用.     

OsmC基因在豌豆根瘤菌抗氧化和共生中的功能

为研究OsmC基因在豌豆根瘤菌抗氧化和共生固氮中的功能,以豌豆根瘤菌RL3841的OsmC基因为研究对象,通过同源重组获得了OsmC基因突变株RLOsmC.结果表明:OsmC基因突变不影响豌豆根瘤菌的自生生长能力,但对无机氧化物H_2O_2和低浓度有机氧化物氢过氧化枯烯(CuOOH)都十分敏感.突变株RLOsmC感染豌豆宿主能形成红色有效根瘤,但是其固氮酶活降低了18.3%.OsmC基因的表达不受H_2O_2的诱导,但在7 d根瘤类菌体中表达量显著上调.表明根瘤菌OsmC基因在共生和抗氧化功能中发挥了重要的作用.     

豌豆根瘤菌RND外排转运基因tpaA的功能研究

Rhizobium. leguminosarum 3841 tpa A基因编码RND家族内膜外排转运蛋白,RND外排泵的过量表达是革兰阴性菌细菌抗生素耐药性形成的重要原因之一.采用基因同源重组构建豌豆根瘤菌tpa A基因突变株RLtpa A,并研究了该基因突变对根瘤菌生长、抗生素耐药性以及共生固氮的影响.结果表明,tpa A基因突变会使根瘤菌在AMS基本培养基生长延迟,但不影响其在TY天然培养基中的正常生长. tpa A基因突变可能抑制了根瘤菌中的RND家族外排泵的过量表达,从而使其抗生素耐药性降低.接种了tpa A基因突变株的豌豆植株能形成红色有效根瘤,但其接种根瘤固氮酶活性降低.实时荧光定量RT-PCR分析表明tpa A基因在根瘤类菌体中的表达显著上调.     

用~(15)N 示踪法测定2,4-D 诱发的小麦和向日葵根瘤的固氮活性

用~(15)N 示踪法定性测定了经2,4-D 诱发的、接种豇豆根瘤菌(R.sp),菜豆根瘤茵(R.phaseoli)等四种根瘤菌和圆褐固氮菌(Az.chroococcum)的小麦(Triticum aesticum)和向日葵(Helianthus annuus)根瘤,结果表明具有固氮活性。     

沙打旺根瘤超微结构研究

目的 对沙打旺根瘤中心组织的超微结构进行研究，并讨论拟菌体的衰老及PHB颗粒的变化。方法 石蜡切片法和超薄切片法。结果 通过对沙打旺根瘤的显微和超微结构观察发现：其根瘤显微结构由4部分组成，由外向内依次为：保护层、皮层、鞘细胞层和中心组织(侵染组织)。在中心组织的侵染细胞中，拟菌体的衰老具有一定的规律，开始拟菌体的周膜和细胞壁变得凹凸不平，但周膜仍几乎较紧地包裹着拟菌体，细胞质染色变浅，纤维状的核物质不明显，并出现大量的β-多羟基丁酸盐(PHB)颗粒，沿着拟菌体的细胞壁分布。后来，拟菌体的周膜开始变得模糊，细胞质凝缩为电子致密的团块状物质，与电子透明的区域相间排列，此时PHB颗粒消失。结论 沙打旺根瘤中心组织的侵染细胞中，拟菌体的衰老具有一定的规律．     

农杆菌转化植物细胞的机理及其基因工程

农杆菌T—DNA能进入植物细胞并稳定地保存在植物染色体中,成为植物细胞新增的一群基因,最终能通过有性世代遗传给子代。这是自然界中存在的原核生物对真核生物进行基因工程的奇妙例证。 根癌农杆菌(Agrobacterium tumefacient)和发根农杆菌(A.rhizogenes)的一些菌株几乎侵染所有受伤后双子叶植物的能力,它们是同属于根瘤菌科的革兰氏阴性菌。它们能分别在宿主植物的感染部位引起冠瘿瘤(Crown gall tumor)和发根瘤(Hairy root tumor),一旦诱导后,冠瘿瘤或发根瘤不需要这些细菌也能增殖。这说明农杆菌的遗传物质能进入宿主细     

某些细菌产生的细胞分裂素的研究初探

用根瘤菌和固氮菌的7个菌株进行实验,以紫云英据瘤菌(Rhizobium astr-agali 1.8)产生细胞分裂素的能力最强。其产生的量与菌体生长情况有关。在菌体生长的对数期,产生细胞分裂素的量随着菌体生长量的增加而增加,到稳定期初,产生细胞分裂素的量可达最大值。之后,发酵液中细胞分裂素的量反而明显减少。此种细胞分裂素的紫外吸收特性与玉米素相类似。     

刺槐根瘤菌优良菌株的初步筛选

从豆科植物根瘤分离获得的８９个根瘤菌株，经刺槐水培和盆栽接种试验，９２－１－１１、８４－２１、９２－１２５三个菌株，不仅在刺槐根部形成的根瘤多且大，根瘤的乙炔还原固氮酶活性每克鲜瘤温育１ｈ乙炔还原为乙烯量接近或超过了１０００μｍｏｌ，超过了三叶草优良根瘤菌株和高效大豆根瘤菌株的固氮水平，对刺槐生长有明显促进作用，生长二个月的植株平均鲜重超过５ｇ，比对照高出８２％以上，故认为这三个菌株为刺槐初筛的优     

根瘤菌在菜豆生长中的作用及接种方法

菜豆的根系有根瘤菌共生,根瘤菌在菜豆生长中起着重要作用。根瘤菌是一种小型球状细菌,在土壤中根瘤菌能逐渐长大,并长出鞭毛,能够运动,根瘤菌进入菜豆的根部形成根瘤。菜豆白出苗后20天,主侧根上开始形成根瘤,根瘤菌在根瘤中利用菜豆提供的营养物质生活,并能吸收空气中的二氧化氮并转化为化合态氮,它所固定的氮素又能为菜豆生长利用。菜豆一生所需氮素,     

不同豌豆品种根瘤菌接种效应的比较研究

通过盆栽试验,选取中豌4号(V1)、中豌6号(V2)、成豌8号(V3)、鄂豌567(V4)、红花青豌豆(V5)和中豌5号(V6)共6个品种为实验材料,进行接种根瘤菌(Accc16103)和不接种处理,比较分析接种根瘤菌对不同豌豆品种种植地的土壤肥力、根瘤发育及其固氮酶活性、豌豆产量构成因素及籽粒品质的影响.结果表明,接种根瘤菌后,中豌5号能显著提高有机质含量,而中豌4号、中豌6号、成豌8号、鄂豌567则能显著提高全氮含量;除成豌8号外,其余品种能显著提高碱解氮含量.中豌4号和红花青豌豆可显著提高有效钾含量,而成豌8号、鄂豌567则能显著提高速效磷含量.接种根瘤菌还可以显著提高豌豆品种的根瘤干重(鄂豌567除外)和固氮酶活性,但对不同豌豆品种单株荚数的影响不明显,中豌6号品种接种可显著提高单荚粒数,而中豌4号、中豌6号和成豌8号在接种根瘤菌后均显著提高百粒重.此外,研究还发现,接种根瘤菌还可以显著提高不同豌豆品种籽粒中粗蛋白质含量和粗纤维含量.     

刺槐根瘤发生的超微结构研究

通过扫描电镜和透射电镜观察刺槐（Ｒｏｂｉｎｉａｐｓｅｕｄｏａｃａｃｉａ）幼瘤的超微结构。刺槐根瘤中侵入线丰富、粗大，内具有高电子密度基质，基质中包埋大量根瘤菌。根瘤菌通过降解侵入线壁和从侵入线无壁泡中释放两种方式进入宿主细胞质     

Deregulation of a Ca2+/calmodulin-dependent kinase leads to spontaneous nodule development

Induced development of a new plant organ in response to rhizobia is the most prominent manifestation of legume root-nodule symbiosis with nitrogen-fixing bacteria. Here we show that the complex root-nodule organogenic programme can be genetically deregulated to trigger de novo nodule formation in the absence of rhizobia or exogenous rhizobial signals. In an ethylmethane sulphonate-induced snf1 (spontaneous nodule formation) mutant of Lotus japonicus, a single amino-acid replacement in a Ca2+/calmodulin-dependent protein kinase (CCaMK) is sufficient to turn fully differentiated root cortical cells into meristematic founder cells of root nodule primordia. These spontaneous nodules are genuine nodules with an ontogeny similar to that of rhizobial-induced root nodules, corroborating previous physiological studies. Using two receptor-deficient genetic backgrounds we provide evidence for a developmentally integrated spontaneous nodulation process that is independent of lipochitin-oligosaccharide signal perception and oscillations in Ca2+ second messenger levels. Our results reveal a key regulatory position of CCaMK upstream of all components required for cell-cycle activation, and a phenotypically divergent series of mutant alleles demonstrates positive and negative regulation of the process.     

Host sanctions and the legume-rhizobium mutualism

Explaining mutualistic cooperation between species remains one of the greatest problems for evolutionary biology. Why do symbionts provide costly services to a host, indirectly benefiting competitors sharing the same individual host? Host monitoring of symbiont performance and the imposition of sanctions on 'cheats' could stabilize mutualism. Here we show that soybeans penalize rhizobia that fail to fix N(2) inside their root nodules. We prevented a normally mutualistic rhizobium strain from cooperating (fixing N(2)) by replacing air with an N(2)-free atmosphere (Ar:O(2)). A series of experiments at three spatial scales (whole plants, half root systems and individual nodules) demonstrated that forcing non-cooperation (analogous to cheating) decreased the reproductive success of rhizobia by about 50%. Non-invasive monitoring implicated decreased O(2) supply as a possible mechanism for sanctions against cheating rhizobia. More generally, such sanctions by one or both partners may be important in stabilizing a wide range of mutualistic symbioses.     

诱导不结瘤豆科树种结瘤固氮的研究

根据国内外有关诱导非豆科作物结瘤固氮的现状,结合作者几年来的工作,对诱导不结瘤豆科树种结瘤固氮的可行性进行了探讨,并报道了作者近年来人工诱导不结瘤的豆科树种—山皂荚(Gledtsia melanacantha)的试验结果。     

大豆根瘤内感染细胞原生质体的分离与活力检测

作者在前人工作的基础上，分离了大豆根瘤中感染细胞的原生质体，分离得到的感染细胞原生质体产量较高。经显微观察和几种细胞化学方法检测的结果表明，原生质体纯度高、破损少、具很强的生活力。对于以其为材料的生化研究具有一定的实际应用价值。     

Amino-acid cycling drives nitrogen fixation in the legume-Rhizobium symbiosis

The biological reduction of atmospheric N2 to ammonium (nitrogen fixation) provides about 65% of the biosphere's available nitrogen. Most of this ammonium is contributed by legume-rhizobia symbioses, which are initiated by the infection of legume hosts by bacteria (rhizobia), resulting in formation of root nodules. Within the nodules, rhizobia are found as bacteroids, which perform the nitrogen fixation: to do this, they obtain sources of carbon and energy from the plant, in the form of dicarboxylic acids. It has been thought that, in return, bacteroids simply provide the plant with ammonium. But here we show that a more complex amino-acid cycle is essential for symbiotic nitrogen fixation by Rhizobium in pea nodules. The plant provides amino acids to the bacteroids, enabling them to shut down their ammonium assimilation. In return, bacteroids act like plant organelles to cycle amino acids back to the plant for asparagine synthesis. The mutual dependence of this exchange prevents the symbiosis being dominated by the plant, and provides a selective pressure for the evolution of mutualism.     

Shoot control of root development and nodulation is mediated by a receptor-like kinase

In legumes, root nodule organogenesis is activated in response to morphogenic lipochitin oligosaccharides that are synthesized by bacteria, commonly known as rhizobia. Successful symbiotic interaction results in the formation of highly specialized organs called root nodules, which provide a unique environment for symbiotic nitrogen fixation. In wild-type plants the number of nodules is regulated by a signalling mechanism integrating environmental and developmental cues to arrest most rhizobial infections within the susceptible zone of the root. Furthermore, a feedback mechanism controls the temporal and spatial susceptibility to infection of the root system. This mechanism is referred to as autoregulation of nodulation, as earlier nodulation events inhibit nodulation of younger root tissues. Lotus japonicus plants homozygous for a mutation in the hypernodulation aberrant root (har1) locus escape this regulation and form an excessive number of nodules. Here we report the molecular cloning and expression analysis of the HAR1 gene and the pea orthologue, Pisum sativum, SYM29. HAR1 encodes a putative serine/threonine receptor kinase, which is required for shoot-controlled regulation of root growth, nodule number, and for nitrate sensitivity of symbiotic development.     

河北省豆科植物根瘤菌资源的初步调查研究

通过对河北省全境豆科植物的结瘤和固氮状况进行调查 ,共采集到 2 4属 5 3种豆科植物的根瘤样品 2 3 5份 ,98.1的被调查豆科植物可自然结瘤 ,96.2的自然瘤可测到固氮酶活性 .其中海边香豌豆 (L athyrus maritimus)、野百合 (Crotalaria sessilif lora)、阴山胡枝子 (L athyrusinschanica)、山岩黄耆 (H edysarum alpinum)、辽西扁苜蓿 (Melilotus ruthenicus)的结瘤固氮情况为首次报道 .     

中国沙棘营养器官结构特征及其与生境关系的研究

对中国沙棘初生结构的分化和次生生长过程进行了观察。发现沙棘根系角质层发达，外被粘质物，外皮层细胞的细胞壁大量栓质化。根瘤的形成与木质部和皮层有密切关系，弗兰克氏菌的侵入刺激皮层形成大量的薄壁细胞，向外突出形成根瘤。茎的皮层外部细胞较小，排列整齐，细胞壁厚，表皮外有很厚的角质层和一定量的蜡质。叶的结构更加体现了抗旱、耐寒植物的特点，栅栏组织发达，排列紧密，海绵组织不发达，细胞间隙较大，气孔深陷，且具有角质层和密而叠置的星状毛及鳞片等。这些均为生活在旱、寒环境里表现出的适应性。     

Internalization of the human immunodeficiency virus does not require the cytoplasmic domain of CD4

Binding of the human immunodeficiency virus (HIV) to infectable host cells, such as B and T lymphocytes, monocytes and colorectal cells, is mediated by a high-affinity interaction between the gp120 component of the viral envelope glycoprotein and the CD4 receptor. Upon binding, it is thought that the second component of the envelope, gp41, mediates fusion between the viral envelope and host cell membranes. However, the early steps of HIV infection have not yet been thoroughly elucidated. Viral entry was first reported to be mediated by pH-dependent receptor-mediated endocytosis; subsequent studies have shown entry to be pH-independent. Although direct fusion of virus to plasma membranes of infected cells has been observed by electron microscopy, it is still formally possible that the infectious path of the virus involves receptor-mediated endocytosis. To gain a better understanding of receptor function in viral entry, we have analysed the ability of several altered or truncated forms of CD4 to serve as effective viral receptors. Our results indicate that domains beyond the HIV-binding region of CD4 are not required for viral infection. Some of the altered forms of CD4 that serve as effective HIV receptors are severely impaired in their ability to be endocytosed. These experiments therefore support the notion that viral fusion to the plasma membrane is sufficient for infection.