根瘤菌属几种同工酶的研究

本文对根瘤菌属12株不同根瘤菌的过氧化物酶,谷氨酸草酰乙酸转氨酶及酯酶同工酶进行了测定,结果发现:所有菌株的过氧化物酶和谷氨酸草酰乙酸转氨酶均呈阴性。酯酶同工酶酶带数在快生型和慢生型根瘤菌之间存在明显的区别,在YMA培养基上,快生型根瘤菌仅有1—2条,迁移率在0.71—0.98之间变化;而慢生型根瘤菌却有4—6条酶带,迁移率在0.15—0.98之间变化,同一菌株在不同碳源上酯酶同工酶酶带数也发生显著变化。     

Rhizobial polysaccharide-degrading enzymes from roots of legumes

The surface structure and chemistry of symbiotic bacteria from the genus Rhizobium are probably important for the outcome of the infection of legume hosts. Exopolysaccharide, capsular polysaccharide, lipopolysaccharide and a low-molecular-weight polysaccharide were isolated from R. trifolii UTC 110-1 and R. leguminosarum UTC 114-5 and partially characterized. No or only minor differences in sugar composition could be found for the corresponding fractions from the two organisms. A general method to measure low activities of polymer-degrading enzymes was developed, and used to determine enzyme activities in root extracts of Trifolium repens L. cv. Lena and Pisum xativiini L. cv. Little Marvel against the isolated rhizobial polysaccharides. An enzyme preparation from T. repens partially degraded all polysaccharides isolated from its symbiont R. trifolii while polysaccharides from R. leguminosarum , symbiont of P. sativum , were degraded to a much lesser extent. Correspondingly, an enzyme preparation from P. sativum degraded all polysaccharides isolated from both its symbiont R. leguminosarum and its non-symbiont R. trifolii. The amount of symbiont polysaccharides degraded was larger than the amount of polysaccharides degraded from the non-symbiont R. trifolii.     

Comparison of geographically distant populations of Rhizobium isolated from root nodules of Phaseolus vulgaris

Seventy-two rhizobial strains were isolated from the root nodules of french beans ( Phaseolus vulgaris ). They were sampled from two geographically distant field populations and 18 additional different sites in France. They were characterized by a) plasmid profiles, (b) RFLP analysis of total cellular DNA using various chromosomal and symbiotic gene probes (including nif H from Rhizobium etli bv. phaseoli ) and c) their ability to nodulate a potential alternative host, L. leucocephala. Over half of the isolates were ascribed to Rhizobium leguminosarum bv. phaseoli on the basis of the hybridization analysis, the possession of multiple copies of nif H and their inability to nodulate L. leucocephala. The remaining isolates belonged to 2 groups which were shown to be genomically distinct from R. leguminosarum bv. phaseoli, R. etli bv. phaseoli and R. tropici. Most members of these two groups shared with R. tropici the ability to nodulate L. leucocephala and, for isolates of only one of these groups, the presence of one copy of nif H. Members of each of the 3 taxa were widely distributed in France and circumstantial evidence of pSym transfer between them was shown. R. leguminosarum bv. phaseoli and one of the two novel groups co-occurred within the two geographically distant populations. Individual genotypes were conserved between them. The finding of a third taxon at various other locations indicated additional diversity among rhizobia nodulating beans.     

Isolation of root hairs from seedlings of Pisum sativum. Identification of root hair specific proteins by in situ labeling

A procedure was developed which allows the large-scale isolation of root hairs from seedlings of Pisum sativum . L. cvs. Kleine Rheinländerin and Rosa Krone. The method may yield up to 50 g fresh weight of root hairs per 3.10⁴ seedlings. In a modified form considerable amounts of root hair material may be harvested, even after incubation of the roots in aqueous solutions. Thus, detailed biochemical studies on the root hair system have become feasible.
The occurrence of specific proteins in membrane fractions of P. sativum root hairs was demonstrated as follows: Incubation of root hairs in situ with 3-azidonaphthalene-2,7-disulfonate – a strongly anionic, photoactivated fluorescent marker – followed by gel electrophoresis of membrane fractions showed the presence of root-hair specific proteins which, since the system was intact, suggests that they are on the outer surface of the cells.     

Role of siderophore in iron uptake in cowpea Rhizobium GN1 (peanut isolate): Possible involvement of iron repressible outer membrane proteins

Abstract Siderophore produced by cowpea Rhizobium GN1 (Peanut isolate) was shown to be involved in iron uptake by this organism. Siderophore enhanced iron uptake in iron-starved cells. SDS-PAGE analysis of the outer membrane proteins showed two iron repressible outer membrane proteins with approximate molecular mass of 80 kDa and 76 kDa. A siderophore non-producing mutant, which was unable to grow on a medium containing synthetic iron chelators unless and until iron was added exogenously in the medium, could use siderophore of the wild-type for iron uptake indicating that the receptor for Fe-siderophore complex was intact in the mutant.     

根瘤菌结瘤因子的结构和功能

结瘤因子是根瘤菌分泌的寡糖,它作为外在信号,诱发宿主植物根部各种生理反应。引起根毛变形,诱导皮层细胞分裂,形成根瘤原基,作者主要就这一早期结瘤过程中结瘤因子的结构和功能作一综述。     

The occurrence of hopanoid lipids in Bradyrhizobium bacteria

Abstract Lipid extraction procedures followed by GLC and GLC-MS analysis were used to investigate the triterpenoid content in Bradyrhizobium and Rhizobium bacteria. Unlike the tested strains of Rhizobium bacteria, a range of triterpenoids e.g., squalene and different classes of hopanoid derivatives were detected in bacteria from all Bradyrhizobium strains investigated (different strains from Bradyrhizobium japonicum, Bradyrhizobium elkanii as well as Bradyrhizobium sp.). Furthermore, related compounds were identified from some hopanoid lipids (e.g., diplopterol) that carried an additional methyl group in their molecular structure. The hopanoid content was high in some strains and accounted for more than 40% of the total lipid fraction (e.g., in strains Bradyrhizobium japonicum USDA 110 and USDA 31), while other strains contained only about a tenth of that amount (e.g., Bradyrhizobium japonicum ATCC 10324 and Bradyrhizobium sp. ( Lupinus ) ATCC 10319).     

大豆根瘤菌对福美双杀菌剂抗性的研究

通过不同大豆根瘤菌对福美双抗性的研究证明:不同大豆根瘤菌自身固有的对福美双的抗性不同,一般可耐50～100μg/ml浓度的福美双。种子拌0.3％福美双同时拌根瘤菌会使种子上的根瘤菌数降低。本文探讨了利用自发突变选育双抗菌种的可能性。试验还证明:对福美双抗性低的菌种产生自发突变抗福美双的机率要多。     

自生条件下根瘤菌的氢酶表达与固氮的联系

在自生条件下,研究了根瘤菌的氢酶与固氮酶的共轭表达(coexpression)。氢酶表达受碳源限制和氧耗速率的调节,并为固氮酶表达条件促进,在固氮酶放氢的诱导下而与固氮酶共轭表达。此外,观察到外源氢支持根瘤菌自生固氮的活性,氢支持效应被葡萄糖酸钠和氧抑制。     

Partial 16S rRNA gene sequence diversity and numerical taxonomy of slow growing pigeonpea (Cajanus cajan L Millsp) nodulating rhizobia

An investigation was carried out to determine the diversity of 30 isolates of slow growing pigeonpea nodulating rhizobia based on variations in partial sequences of the 16S rRNA gene and numerical analysis of 80 phenotypic traits. Phylogenetic analysis using molecular sequences of 23 isolates showed that ARPE1 separated from the other isolates at an average distance of >14% divergence level. The other isolates were all within 5% divergence from each other but separated into four main groups, with group 1 containing 16 of the 23 isolates. Comparisons to sequences of reference strains revealed that the group 1 isolates were phylogenetically closely related to the slow growing soybean nodulating rhizobia belonging to Bradyrhizobium elkanii, although only three of these isolates were able to nodulate soybean. Numerical analysis of phenotypic data of 19 isolates showed that 14 isolates clustered together in one branch of the phenogram, which included the group 1, group 2 and group 4 isolates from the phylogenetic analysis. The group 3 isolates were highly variable in the phenogram with similarity levels lower than 50% among these isolates.     

环境因子对根瘤菌吸氢活性表达的影响

自生条件下,测定准噶尔盆地南部的68株根瘤菌吸氢活力,获得一株Hup~+的冬箭筈豌豆根瘤菌C_(48)。经与紫云英根瘤菌株109及89比较,两者氢酶表达的最适pH相同;温度分别以20或30℃为宜;Ni~(2+)显著促进吸氢表达,但C_(48)还受Co~(2+ )、Mg~(2+)、Cu~(2+)的促进;紫云英根瘤菌的氨酶表达受碳水化合物抑制较冬箭筈豌豆根瘤菌明显。此外,自生条件下生长的Hup—菌株,经与宿主共生后,Hup~+的百分率大为增加。     

Transconjugants of Agrobacterium radiobacter harbouring sym genes of Rhizobium galegae can form an effective symbiosis with Medicago sativa

It is known that the Rhizobium galegae genomes contain megaplasmids. The suicide vector pSUP2111 with nifH gene of R. meliloti was introduced into the strains CIAM 0703 and CIAM 0711 of R. galegae inducing effective nodules on Galega orientalis plants. The formation of self-transmissible megaplasmids was observed. The megaplasmid transfer into non-nodulating R. meliloti mutants resulted in partial complementation of the nodulation defect in recipient strains though only one transconjugant showed the nitrogen-fixing activity in symbiosis with alfalfa and another one in symbiosis with G. orientalis plants. Among the Agrobacterium strains harbouring R. galegae megaplasmids there were four classes of transconjugants: (1) Nod+ Fix- in symbiosis with goat's rue plants (three strains); (2) Nod+ Fix- on Medicago sativa (two strains); (3) Nod+ Fix+ on M. sativa (five strains); (4) Nod- with both plant hosts (11 strains).     

根瘤菌选育研究进展

生物固氮是一个全球性的战略课题,其中豆科植物与根瘤菌共生固氮一直是生物固氮研究的焦点。该文从菌株选育的角度,通过对比总结国内外根瘤菌选育方法的研究进展,详细阐述了各种育种方法在根瘤菌选育过程中的应用和优缺点,指出筛选周期过长和筛选技术低效是当前研究中的限制问题,并进一步对选育工作的前景进行了展望。     

异叶银合欢根瘤菌的分离和特性

从异叶银合欢K(164)和K(794)的根瘤中各分离到一株根瘤菌,并且,对K164根瘤菌的形态特征和生理生化特性进行了研究。K(164)根瘤菌在平板上培养2天,菌落呈圆形,灰白色,直径约1．2mm;革兰氏染色呈阴性,无芽孢,有美膜,两根极生鞭毛;杆菌大小为2．2×0．89μm,增代时间为1．69小时,属快生型;最适生长温度为30℃,在39℃持续2天不能存活,在20℃以下不能生长;不耐盐,较耐酸,在pH4.5～6．6范围内均能生长,但不耐碱;对庆大霉素和青霉素敏感;对供试的10种糖都能利用,不液化明胶,不利用淀粉,B．T．B试验未变色,不能进行3-酮基乳糖反应,硝酸盐反应呈阳性,石蕊牛奶试验能胨化、还原、产碱,牛肉膏蛋白胨培养生长差,能利用柠檬酸盐;在自生条件下具有吸H2酶活性;回接后结瘤率达100％,根瘤具有固氮酶和吸H2酶活性。     

In vivo expression technology (IVET) selection of genes of Rhizobium leguminosarum biovar viciae A34 expressed in the rhizosphere

IVET was used to identify genes that are specifically expressed in the rhizosphere of the pea-nodulating bacterium Rhizobium leguminosarum A34. A library of R. leguminosarum A34 cloned in the integration vector pIE1, with inserts upstream of a promoter-less purN:gfp:gusA, was conjugated into purN host RU2249 and recombined into the genome. After removal of colonies that expressed the reporter genes of the vector under laboratory conditions, the library was inoculated into a nonsterile pea rhizosphere. The key result is that 29 rhizosphere-induced loci were identified. Sequence analysis of these clones showed that a wide variety of R. leguminosarum A34 genes are expressed specifically in the rhizosphere including those encoding proteins involved in environmental sensing, control of gene expression, metabolic reactions and membrane transport. These genes are likely to be important for survival and colonization of the pea rhizosphere.     

根瘤菌属胞外多糖的研究

本文研究了豌豆根瘤菌(Rhizobum Leguminosarum),苜蓿根瘤菌(R. meliloti),三叶草根瘤菌(R. trifolii),菜豆根瘤菌(R. phaseoli),豇豆根瘤菌(Rradyrhizobium sp.(Vigna))和大豆根瘤菌(R. Japonicum)产生的胞外多糖化学组分的差异,结果表明,不同种的根瘤菌能产生具有不同组分的胞外多糖,其多糖组分的差异主要表现在糖醛酸和甘露糖的含量。豌豆根瘤菌、三叶草根瘤菌,菜豆根瘤菌产生的胞外多糖含有糖醛酸,大豆根瘤菌和苜蓿根瘤菌产生的胞外多糖一般不含有糖醛酸。根瘤菌有快生型和慢生型之别,这种差异也可由其产生的胞外多糖组分看到,一般快生型根瘤菌:豌豆根瘤菌,苜蓿根瘤菌,菜豆根瘤菌,三叶草根瘤菌,(包括最近证明的快生型大豆根瘤菌)的胞外多糖中甘露糖所占百分比较低(低于20％),葡萄糖所占的百分比较高(高于60％),而慢生型根瘤菌:大豆根瘤菌和豇豆根瘤菌的胞外多糖中甘露糖所占百分比较高(高于36％),葡萄糖所占的百分比较低(低于50％)。     

Chemotaxis in Rhizobium meliloti strain L5.30

We have found that Rhizobium meliloti strain L5.30 exhibits positive chemotaxis towards some amino acids, sugars, exudates and extracts from roots of legume plants. From the investigated compounds sugars were better chemo-attractants than amino acids, but legume root substances were the best ones. Positive chemotaxis towards legume root compounds was supported by clouds of R. meliloti cells observed at the surface of alfalfa roots. A large deletion, in the nodABC region of the symbiotic megaplasmid (Sym), did not eliminate rhizobial chemotaxis.     

大豆根瘤中多磷酸盐积累的特征

我们用透射电镜观察了丰收11号大豆根瘤中多磷酸盐颗粒的形态结构、分布规律以及与根瘤菌发育的关系。观察表明,它是一种电子密度很高的圆形颗粒,其直径在110—140nm之间。表面圆整,没有膜包围,内部质地均匀而致密。它只存在于根瘤菌的DHA纤维上,而不存在于胞质中。在刚从侵入线释放出来的根瘤菌中没有或只有很少这种颗粒,但在即将成熟的根瘤菌中却非常丰富,随后又逐渐减少,乃至完全消失。由此可见,根瘤中多磷酸盐的积累与根瘤菌的生长发育有关。