豆科植物-根瘤菌共生固氮的分子机理

与豆科植物－根瘤菌共生固氮有关的基因涉及根瘤菌基因和宿主基因，根瘤菌基因有结瘤基因（nodD,nodAB-CIJ和hsn基因），根瘤菌细胞表面结构基因（exs,lps和ndv基因）和固氮基因（nif和fix基因）；宿主基因主要是结瘤素基因（ENOD和NOD基因）。根瘤菌结瘤基因表达后诱导产生结瘤因子。在根瘤发育过程中，这些基因在根瘤菌与植物之间进行着信息交换，并且具有不同的表达水平。结瘤因子和植物激素对它们进行着调节。     

华癸中生根瘤菌脂多糖突变株的共生能力

通过植物盆栽实验,对华癸中生根瘤菌（Mesorhizoblum huakuii）lpsH基因的缺失突变株HK241进行共生能力研究．结果表明：突变株HK241形成的根瘤没有固氮能力,同时具有较低的结瘤效率和竞争结瘤能力．因此,lpsH基因参与编码脂多糖的合成,并且在共生中发挥非常重要的作用．     

华癸中生根瘤茵脂多糖突变株的共生能力

通过植物盆栽实验,对华癸中生根瘤菌(Mesorhizobium huakuii)lpsH基因的缺失突变株HK241进行共生能力研究.结果表明:突变株HK241形成的根瘤没有固氮能力,同时具有较低的结瘤效率和竞争结瘤能力.因此,lpsH基因参与编码脂多糖的合成,并且在共生中发挥非常重要的作用.     

谈小高炉结瘤的处理

研究了唐山松汀钢铁公司 3#高炉结瘤的原因 ,找出了解决的办法 ,取得了良好效果。     

类根瘤对烟草烟碱含量的影响

对人工诱导结瘤后不同培养时期烟草根、茎、叶柄和叶中的烟碱、灰分和水分含量分别进行了测定。测定结果表明：烟碱和灰分含量，不仅与材料有关，也受培养时间的影响，并随培养时间的增长而增加，与此相反，水分在培养过程中则呈现出一种下降趋势。     

根瘤化对烟草不同叶位叶片中糖的影响

作者对人工诱导结瘤烟草和未结瘤烟草不同叶位叶片中的总糖及可溶性糖、淀粉和粗纤维进行了测定．结果表明：它们的含量都随叶位不同而并，前者高于后者，但下叶位叶片中的粗纤维含量例外。这种分布可能与叶片细胞的生理状态和结瘤有关。     

花生根瘤菌诱变株感染大豆结瘤和固氮

花生根瘤菌诱变株感染大豆结瘤和固氮龙敏南，许良树，张凤章，曾定（生物学系）根瘤菌在共生固氮过程中会放Ｈ２，放Ｈ２是种能量浪费．Ｅｖａｎｓ等［１］研究表明：利用具有吸氢酶活性的根瘤菌［ＨＵｐ＋］接种豆科作物能提高固氮效率、增加作物产量．豆科植物根瘤菌的...     

Isolation and characterization of a <Emphasis Type="Italic">Sinorhizobium fredii</Emphasis> mutant that cannot utilize proline as the sole carbon and nitrogen source

Sinorhizobium fredii strain HN01 can use proline as the sole carbon and nitrogen source. A mutant strain GXHN100 unable to catabolize proline was screened from 6000 Tn5gusA5 random insertional mutants of S.fredii strain HN01. Sequencing analysis showed that an open reading frame, named pmrA (proline metabolic relative), was inserted by the Tn5gusA5. A positive clone, namedp GXHN100 which containing 3.3kb foreign DNA fragment of S.fredii strain HN01, was isolated from a partial gene library of S.fredii HN01 by colony in situ hybridization. Sequence analysis showed that pGXHN100 contained the entire pmrA gene. The 3.3kb DNA fragment of pGXHN100 was cloned into a broad-host-range cosmid vector pLAFR3 to form plasmid pGXHN200 which was subsequently introduced into GXHN100 to form a complemented strain GXHN200. Plant test showed that GXHN100 was effective and no obvious changes in nitrogenase activity comparing with parental strain. But GXHN100 nodulated 2 days later on soybean and its nodulation efficiency and competitiveness were decreased.The complemented strain GXHN200 restored the nodulation efficiency and competitiveness of GXHN100 to the wild type.     

Extra-copy nifA enhances the nodulation efficiency of Sinorhizobium fredii

Previous investigations have shown that nifA gene is involved in nodulation and symbiotic nitrogen fixation regulation of Rhizobium. We study the role of nifA on nodulation of leguminous plants. We found that Sinorhizobium fredii harboring multi-copy plasmid carrying the constitutively expressed Klebsiella pneumoniae nifA exhibited an increase of noduiation activity and nodulation competitiveness on soybean plants. The Nod-factor secreted by the rhizobia cells containing the multi-copied nifA was assayed,and preliminary results showed that S. fredii containing the multi-copy plasmid carrying nifA produced higher strength of Nod-factor than the rhizobia containing the same plasmid carrying the vector did.     

Sinorhizobium meliloti nifA gene exerts a pleiotropic effect on nodulation through the enhanced plant defense response

Sinorhizobium meliloti nifA gene is required for the expression of a bunch of nif and fix genes. Here, we report its pleiotropic effects on the nodule formation. Compared with wild type strain, nifA mutant sig- nificantly reduced nodule suppression rate in split-root system. The plants inoculated with mutant strain produced lower amount of daidzein and less necrotic cells on their roots. In addition, the defense genes failed to be evoked by nifA mutant at the early nodulation stage. These findings indicated that host defense response was one of the mechanisms mediated by nifA gene to regulate nodule formation during symbiosis. Even though nifA mutant could increase the number of nodules in host plant, it synthesized lower Nod factors than wild type. This suggested that nifA gene mediated multiple and diverse instances in nodulation formation.     

两株花生根瘤菌的比较研究及结瘤能力的测定

从花生根瘤中筛选出两株生长速度差异明显的花生根瘤菌:1K(快生型),2M(慢生型),通过16S rDNA测序分析并结合生理生化实验确定两种根瘤菌分别为中华根瘤菌和慢生根瘤菌。对二者的生理特性、抗逆性和结瘤能力等指标进行了研究,发现慢生型虽然在生长速度、抗逆性等方面明显不及快生型,但是结瘤能力比快生型要强。     

A proteomic analysis of bacterial strain<Emphasis Type="Italic">Sinorhizobium fredii</Emphasis> RT19 subjected to salt shock

Sinorhizobium fredii RT19, a strain of freeliving bacteria, was subjected to salt shock and its protein expression profiles were analyzed by differential display proteome approaches. The results of separation by two-dimensional polyacrylamide gel electrophoresis (2D PAGE) showed that the number of resolved proteins was 481, 465 and 424, corresponding to salt-free control, 5 and 50 min 1 mol/L salt treatment, respectively. Among the resolved proteins, 82 in total had altered expression in response to salt-shock stress. 26 out of the 82 proteins were induced and 23 were completely inhibited, while 12 were up-regulated and 21 down-regulated in response to salt shock. In addition, the appearance of differentially displayed proteins responding to different salt shock periods is also reported. The identity of the 26 induced proteins was revealed by matrix assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF/MS) followed by database searching. Among them, 20 were assigned to proteins with known functions. Their roles in response to salt shock stress are discussed.     

慢生型大豆根瘤菌染色体的16kb EcoRⅠ DNA片段的亚克隆测序分析

亚克隆测序分析发现，在Bradyrhizobium japonicum的GX201菌株染色体的16kb EcoR I DNA区段上含有一个与putA基因同源的基因的ORF 3054（Open Reading Frame)，该基因ORF长3054bp，在核苷酸水平上与已报道的putA基因有94％一致性，其推断性的编码产物在氨基酸水平上与putA编码产物脯氨酸脱氢酶（ProDH)有95％一致性，利用Tn5gusA5诱变的方法获得了该同源基因的标记置换突变体，该突变体在液体培养基（YMA）中的生长速率比野生菌株慢。     

Present status and development on biological nitrogen fixation research in China

This presentation introduces the advances in biological nitrogen fixation research abroad, in particular,describes the great progress and achievements on its research in China as follows: collection of rhizobial resources and establishment of the largest database of Rhizobium in China, correction and development of Rhizobium taxonomy in international; discovery of a couple of nif genes, identification and unification of linkage among the nif gene operons of Klebsiella pneumoniae, finding of regulative mechanism of positive regulation nif gene and its sensitivity to oxygen,temperature; finding of the activity of nodulation gene nodD3 product in Sinorhizobium meliloti which is not controlled by flavonoid produced from its host alfalfa; finding of the association between expression of genes coding the products for carbon utilization and nitrogen metabolism and their regulations; chemical synthesis of nodulation factor of Sinorhizobium meliloti; constructions of engineered nitrogen fixers and utilization in practice based on the research of gene expression and regulation; chemical simulation of the structure and function of nitrogenase and bringing forward the model of nitrogenase active center for the first time in international and synthesis of model compounds which were paid attention by colleagues abroad. Finally, the development of nitrogen fixation research in China in future has been put forward, suggesting that the nif gene regulation and its role in providing crops with nitrogen element, signal transduction and molecular interactions between Rhizobium and legume, coupling between carbon and nitrogen metabolisms, nitrogen fixation and photosynthesis, and functional genomics of nitrogen-fixing nodule symbiosis, etc., would be actively worked on.     

鸡新城疫病毒HN蛋白多表位抗原的表达与免疫原性研究

有效抗原及表位的预测和筛选是疫苗研究的基础,在对鸡新城疫病毒HN蛋白抗原表位预测的基础上,对多表位抗原进行表达与免疫原性测定。根据生物信息学表位预测方法获得的家禽新城疫病毒抗原表位,利用PCR技术合成基因,构建pBVIL1-HN重组载体,转化大肠杆菌HB101,进行基因工程表达;经纯化蛋白后免疫小鼠,抗体滴度用酶联免疫吸附方法测定,确定抗原的免疫原活性。结果表明,多表位抗原基因经测序结果正确,融合基因在大肠杆菌得到高效表达,电泳纯融合多表位抗原经三次免疫得到抗血清,抗体滴度为1：8000。鸡新城疫病毒HN蛋白多表位抗原得到高效表达,且具有良好的免疫原性。     

Sinorhizobium meliloti lsrB is involved in alfalfa root nodule development and nitrogen-fixing bacteroid differentiation

Rhizobia interact with host legumes to induce the formation of nitrogen-fixing nodules, which is very important in agriculture and ecology. The development of nitrogen-fixing nodules is stringently regulated by host plants and rhizobial symbionts. In our previous work, a new Sinorhizobium meliloti LysR regulator gene (lsrB) was identified to be essential for alfalfa nodulation. However, how this gene is involved in alfalfa nodulation was not yet understood. Here, we found that this gene was associated with prevention of premature nodule senescence and abortive bacteroid formation. Heterogeneous deficient alfalfa root nodules were induced by the in-frame deletion mutant of lsrB (lsrB1-2), which was similar to the plasmid-insertion mutant, lsrB1. Irregular senescence zones earlier appeared in these nodules where bacteroid differentiation was blocked at different stages from microscopy observations. Interestingly, oxidative bursts were observed in these nodules by DAB staining. The decreased expression of lipopolysaccharide core genes (lpsCDE) was correspondingly determined in these nodules. S. meliloti lipopolysaccharide is required for suppression of oxidative bursts or host cell defense. These findings demonstrate that the S. meliloti lsrB gene is involved in alfalfa root nodule development and bacteroid differentiation by suppressing oxidative bursts or defense responses in host cells.     

黄河三角洲地区野大豆根瘤菌生物学特性及其遗传多样性研究

野大豆具有耐盐碱、抗寒、抗病等特点,对于研究大豆遗传基因的变迁、改善大豆品质和产量具有重要作用。为了探明黄河三角洲地区野大豆根瘤菌的生物学特性及其遗传多样性,从黄河三角洲地区的14个乡镇26个采样点采集了227株野大豆,从其根瘤中分离纯化根瘤菌,进行一系列的生理生化实验、抗逆性实验、结瘤能力测定、结瘤广谱性测定、16S rDNA序列测定和RAPD分析,共分离纯化出100 株根瘤菌,其中菌株3D-21,3D-24,3K-8,3K-23VS等对酸、碱、盐、抗生素、高温、低温均有较强耐受性,且结瘤能力也较强;菌株7K-8结瘤能力较强,K-5抗逆性较强。16S rDNA序列测定表明所获菌株分属于3个属7个种,相近菌株的RAPD分析呈现明显的多态性。结果表明黄河三角洲地区野大豆根瘤菌存在着丰富的多样性,部分菌株结瘤能力和/或抗逆能力强,该研究为挖掘、利用优良菌株资源奠定了基础。