Assessment of nifH diversity in rhizobial isolates of different origin and the role of antioxidant in respiratory protection

Rhizobia diversity is considered as one of the most useful resources for bioprospecting due to their symbiotic nitrogen-fixing ability with members of Leguminosae. The highly conserved nature of the nitrogenase reductase gene (nifH) makes it an ideal molecular tool to determine the potential for biological nitrogen fixation in any environment. In the present investigation, 250 rhizobial strains were isolated from legumes belonging to different geographical locations of Chhattisgarh, India. Genetic diversity of the nitrogenfixing bacterial community was analyzed using the nifH gene-specific primer. The polymorphism was found among the nitrogen-fixing population of different sources and origin but not in same source of rhizobia. Further, the symbiotic plasmid DNA was characterized on the basis of size and copy number of plasmids. The plasmid number varying from one to three in different rhizobial isolates had a size greater than 23 kb, while in some rhizobial isolates plasmids were absent. In addition, to examine the role of ascorbate in respiratory protection, the clear black spot margin of ascorbate was observed in the endodermis region of the nodule whereas scarcely dispersed in the infected region. Therefore, our findings demonstrated that knowing the rhizobial nifH gene diversity along with copy number of the plasmid is important for strain identification, deciding its fertility, productivity standards, and potential of biological nitrogen fixation across the geographical region.     

Isolation and genetic characterization of a fertility-restoring revertant induced from cytoplasmic male sterile rice

Summary A male fertile revertant was isolated from M₁ of a cytoplasmic male sterile indica rice line II-32A, the dry seeds of which were treated with ⁶⁰Co- rays at a dose of 290 Gy. The acquired revertant T24 was morphologically and agronomically similar to II-32B, the maintainer of II-32A. Testcrosses of the revertant with II-32A and Zhenshan 97A showed that the revertant was able to restore the fertility of CMS lines. Genetic analysis of the progenies between T24 and II-32A, Zhenshan 97A XieqingzaoA and DZhenshan 97A, which have different cytoplasms, showed that the fertility restoration of four CMS lines by T24 involved one nuclear gene, indicating that T24 was a result of the mutation of one nuclear gene. The mechanism of the restoration of CMS line by T24 was obviously different from other restorers such as Minghui 63 and 20964, which were shown to behave in two gene mode in fertility restoration. The discovery of the revertant T24 contributes to the understanding of CMS and fertility restoration of CMS in rice. The T24 and its parent II-32A may constitute a pair of near isogenic lines for the restoring gene, which should be valuable materials for molecular genetic analysis of CMS.     

A型产气荚膜梭菌的分离及鉴定

从山西各鹿场采集鹿出血性肠炎急性病死鹿病料39例,通过病原微生物分离培养,形态学观察和血清型鉴定,初步表明分离到的产气荚膜梭菌主要是A型,对 PCR产物的序列分析表明经过PCR得到了特异性的α毒素基因片段,因而A型产气荚膜梭菌是引起山西鹿场发生鹿出血性肠炎的主要致病菌。为该疾病的防治和疫苗研制提供依据。     

Isolation and linkage mapping of disease-resistance-like sequences from various rice cultivars, containing different recognition specificities

Degenerated oligonucleotide primers identified from the nucleotide‐binding sites of known disease resistance (R) genes were used from rice cultivars harbouring different recognition specificities to amplify and clone homologous sequences of R genes. A total of 68 non‐redundant clones, which showed various degrees of sequence homology to R genes, were obtained from 18 rice cultivars. These clones had a high degree of sequence diversity both in the nucleotides and in the predicted amino acids, and were classified into five groups using clustal analysis. Fifteen of the 68 clones were mapped to 17 loci on chromosomes 3, 5, 11 and 12 in the rice molecular linkage map. The loci of the mapped clones correlated with the locations of known rice R genes for blast resistance and bacterial blight resistance on chromosomes 11 and 12. Other mapped loci occurred in cluster on chromosome 3, and correlated with the position of a quantitative trait locus for bacterial blight resistance. The mapping of the R gene homologues may aid the identification and isolation of R gene candidates.     

Isolation and characterization of microsatellite markers from guineagrass (Panicum maximum) for genetic diversity estimate and cross-species amplification

Guineagrass ( Panicum maximum Jacq.) is one of the major forage grasses in tropical and semitropical regions, largely apomictic and predominantly exist as tetraploid. Non-availability of polymorphic molecular markers has been a major limitation in its characterization and improvement. We report isolation and characterization of microsatellites in P. maximum and cross-species results with other five Panicum species. Based on microsatellite-motifs, 15 functional and polymorphic simple sequence repeat (SSR) primer-pairs were designed, validated and employed in estimating genetic relationship among 34 guineagrass accessions. Thirteen primer-pairs amplified single locus and remaining two generated more than two loci with an average of 3.57 bands per locus amounts to 63 bands with 34 guineagrass accessions. Average expected heterozygosity ( H _E) of 0.35 (maximum 0.97) and observed heterozygosity ( H _O) of 0.37 (maximum 0.91) established the efficiency of developed markers for discriminating guineagrass accessions. Dice's similarity coefficients-based unweighted pair group with arithmetic average method-clustering supported with high bootstrap values (≥40) indicated its significance and distinguished all accessions except IG97-93 and IG97-6. Utility of these new SSR loci in genetic diversity study of P. maximum and other cross–amplified species is discussed.     

Isolation and characterization of resistance gene analogs (RGAs) from sorghum (Sorghum bicolor L. Moench)

Degenerate primers designed based on known resistant genes (R-genes) and resistance gene analogs (RGAs) were used in combinations to elucidate RGAs from Sorghum bicolor, cultivar M 35-1. Most of the previously tried primer combinations resulted in amplicons of expected 500–600 bp sizes in sorghum along with few novel combinations. Restriction analysis of PCR amplicons of expected size revealed a group of fragments present in a single band indicating the heterogeneous nature of the amplicon. Many of these were cloned and some were considered for analysis. The nucleotide sequence of different cloned fragments was done and their predicted amino acid sequences compared to each other and to the amino acid sequences of known R-genes revealed significant sequence similarity. A cluster analysis based on neighbor-joining (N-J) method was carried out using sorghum RGAs (SRGAs) together with several analogous known R-genes resulting in two major groups; cluster-I comprising only SRGAs and cluster-II comprised of known R-gene sequences along with three SRGAs. Further analysis clearly indicated similarity of SRGAs in overall sense with already known ones from other crop plants. These sequences can be used as guidelines to detect, map and eventually isolate numerous R-genes in sorghum.     

巴西大豆中炭疽菌的分离鉴定研究

此文中通过对巴西进境大豆病害分离,共得到30个菌株,并对其中的4株炭疽菌进行了形态学和分子生物学鉴定,确认了它们分别是胶孢炭疽菌（Colletotrichum gloeosporioide）、辣椒炭疽菌（Colletotrichum capsici）、平头炭疽菌（Colletotrichum truncatum）和黑线炭疽菌（Colletotrichum dematium）。此研究从30个菌株中分离得到的4株炭疽菌分属于不同种,从而证实了巴西大豆中炭疽菌的多样性,不仅充实了正在构建的大豆病害数据库和菌种资源库,也可为港口的植物检疫工作提供借鉴。     

沙棘内生菌的分离与初步鉴定

植物内生菌广泛存在于各种植物中,在与宿主的协同进化过程中形成了两者的互惠共生关系。通过对采集的野生沙棘健康的根、茎、叶进行组织分离共计获得内生真菌327株,分别归属于链格孢属、青霉属、根霉属、芽枝霉属、毛壳菌属、曲霉菌属、盘多毛孢属、向基孢属、枝顶孢属、头孢霉属等10个真菌属,内生菌表现出丰富生物多样性。从组织分离以及对内生真菌初步鉴定的结果看一些物种还具有组织专化性。     

婴儿粪便中益生菌的分离筛选及培养

采用从婴儿粪便中提取具有益生作用的菌种,为下一步的研究开发做乳酸菌种库做积累。选取河南省洛阳市5位志愿者进行样品采集,得到5份完全由母乳喂养的婴儿(2月龄)自然排出的粪便样,并分离纯化出5株乳酸菌(RS1, RS2,RS3,RS4,RS5)和5株双歧杆菌(SQ1,SQ2,SQ3,SQ4,SQ5),经鉴定,确定RS3和SQ1具有益生作用的。结果表明,此2株菌具有较好的耐酸性、胆盐耐受力,对温度敏感,对模拟胃液和肠液具有较强的耐受力,且对大肠杆菌、沙门氏菌具有95%以上的抑菌率,可作为优良的益生菌候选菌种。     

我国梨和部分国外梨果实上链格孢菌的鉴定研究

本文从链格孢的产孢表型、分生孢子特征和分子生物学方面研究了从河北鸭梨、新疆库尔勒香梨、美国和日本以及智利的梨上分离的链格孢菌的种类.经对115支Alternaria菌株的形态学鉴定,共确定了3个种,即Alternaria alternata、A.tenuissima和A.infectoria.ITS系统发育分析研究表明,Alternaria大孢子种彼此间及大孢子种与小孢子种间已发生明显分化,可以根据ITS序列差异明确区分.而供试的多数小孢子种在ITS序列上差异很小,因而不能根据ITS序列差异来区分.     

从植物中分离苏云金芽孢杆菌的研究

苏云金芽孢杆菌是目前研究最深入、应用最广泛的杀虫微生物,广泛分布于世界各地。从热带雨林到北极冻土带,从昆虫尸体、野生动物、动物粪便、贮藏物或尘埃、植物表面、鲜水到土壤中都有Bt存在。以植物叶片作为Bt菌株分离资源的相关研究相对较少,而非维管束植物中分离Bt菌株的报道更是屈指可数。从植物中分离Bt不但能丰富Bt的生态学意义,还能极大地丰富Bt的菌种资源。本文就植物上分离微生物、维管植物分离Bt的研究进展及非维管植物分离Bt的研究前景做简单介绍。     

Isolation and characterization of microsatellites in Bambusa arundinacea and cross species amplification in other bamboos

Isolation and characterization of microsatellites was analysed in Bambusa arundinacea and cross species amplification studied in other bamboos. Microsatellites, tandem repeats of short nucleotide (1–6 bp) sequences, are the DNA marker of choice because of their highly polymorphic, ubiquitous distribution within the genome, ease of genotyping through Polymerase chain reaction, selectively neutral, co‐dominant and multiallelic nature. Six microsatellites, three polymorphic and three monomorphic have been characterized for the first time in a bamboo species, Bambusa arundinacea belonging to the family Poaceae. The numbers of alleles per locus ranged from 2 to 13. Cross species amplification was tested in 18 other bamboo species. Monomorphic simple sequence repeats (SSRs) were found to be cross amplified in most of the species tested and polymorphic ones in only three to four species. The utility of SSR loci in a genetic diversity study of B. arundinacea and other cross‐amplified bamboo species has been discussed. This study will help in population genetic studies in bamboo species.     

Isolation and characterization of a rice mutant insensitive to cool temperatures on amylose synthesis

The Wx ^b gene, one of the alleles at the rice waxy(wx) locus, is activated at cool temperatures during seed development, andas a result, a large amount of amylose is accumulated causing a reductionin rice grain quality. We found that the seeds of a du mutant couldbe visibly distinguished depending on whether they matured at cool ornormal temperatures. Using these characteristics, we isolated a mutantcandidate insensitive to cool temperatures. While the amylose content inthe original line was about 2% at a normal temperature (28 °C)and 12% at a cool temperature (21 °C), in the mutant candidate(coi) the amylose content was not affected by temperatures, i.e. theamylose content was about 3% at both temperatures. This finding incombination with the results of an immunoblot analysis indicated that theabsence of an increase in the amylose content in this mutant was caused bya constant level of Wx gene expression at normal and cooltemperature. Genetic analysis revealed that this insensitivity to cooltemperatures was caused by a single recessive mutation. This mutantshould be useful in breeding programs designed to produce rice of desiredquality at cool temperatures and in understanding genetic and molecularmechanisms that respond to slight changes in temperature.     

Analysis of host genotype x rhizobial strain interaction in N2 fixation

Summary A method of analyzing host genotype x rhizobial strain interaction for N₂ fixation potential, based on the principle of structural relationship analysis, is proposed. When this interaction is significant, selecting genotypes for high N₂ fixation potential is incorrect, because the N₂ fixation efficiency is dependent on the rhizobial strain used. Under such circumstances, grouping genotypes based on average fixing ability (AFA), a linear response to the effectiveness of rhizobial strain, and specific fixing ability (SFA), deviation from the linear response in terms of the magnitude of the error variance, is useful before initiating a breeding program for enhanced N₂ fixation. Host genotype x strain interaction effect is partitioned into two components, and two parameters are derived which estimate AFA and SFA. N₂ fixation data from 3 mungbean genotypes and 6 rhizobial strains are used to illustrate this method.     

Isolation, identification and characterization of disomic and translocated barley chromosome addition lines of common wheat

Two disomic barley chromosome addition lines and five translocated chromosome addition lines of common wheat cultivar Shinchunaga were isolated. They were derived from a hybrid plant between Shinchunaga and cultivated barley Nyugoruden (New Golden) by backcrossing with wheat and self pollination. Barley chromosomes added to chromosome arms involved in the translocated chromosomes were identified by C-banding method and by crossing these lines with Chinese Spring/Betzes addition lines. Two disomic addition lines were identified to have chromosome 6 and 7 of barley, respectively. Two of the five translocated chromosome addition lines were clarified to have same chromosome constitution, 42 wheat chromosomes and a pair of translocated chromosomes constituted with a long arm of chromosome 5B of wheat and a short arm of chromosome 7 of barley. The other three lines could not be identified due to chromosome rearrangement. Performances of these seven lines on agronomic characters were examined. Addition of barley chromosome 7 induced early heading, and chromosome 6 showed lated heading. Almost all of the lines except that of chromosome 6 showed short culm length and all showed reduced number of tillers, spikelets and grains per ear, and low seed fertility. These lines would be useful for genetic analyses in wheat and barley and for induction of useful genes of barley into wheat. This revised version was published online in July 2006 with corrections to the Cover Date.