阎氏菌科的16S rRNA可变区二级结构分析

采用16SrRNA可变区二级结构图形分析,比较了阎氏菌科阎氏菌属典型种与微球菌亚目中几个相关科属典型种可变区二级结构的变化。结果表明,V3、V4存在明显的不同。将16SrRNA二级结构划分成不同的结构单元,提出在9个可变区中,至少要有2个不同的结构单元才可以定为新科,存在1个不同的结构单元可以定位新属;并认为16SrRNA可变区二级结构分析,可以作为一种辅助手段,应用于原核生物属以上水平的分类。     

16S rRNA analysis and the phylogenetic position of Wolinella succinogenes

The 16S ribosomal RNA of Wolinella succinogenes ATCC29543 was analyzed by the RNase T1 oligonucleotide cataloguing approach. In contrast to its present classification, W. succinogenes is related neither to members of the genus Bacteroides nor to any other genus of the family Bacteroidaceae. As derived from the similarity coefficients (S_AB values) calculated on the basis of more than 350 eubacterial species, W. succinogenes appears to be a distantly related member of the division of purple photosynthetic bacteria and their relatives; however, S_AB values do not indicate that this species is preferentially related to any representative of the 4 subdivisions.     

基于线粒体16S rRNA基因对中国狼蛛科(Lycosidae)系统发育研究

将自测的中国狼蛛科Lycosidae4亚科6属26种和从GenBank中检索到的北美2种豹蛛的mtD-NA-16S rRNA序列进行比较;以漏斗蛛科1种蜘蛛作为外群,对碱基序列的组成和遗传距离进行了分析,采用Bayesian方法和最大简约法(MP)构建分子系统树。研究结果表明:16SrRNA基因的部分序列为340bp到360bp,A T含量平均为75%,存在较强的A T含量偏向性;序列共有157个碱基存在变异,其中79个简约信息位点。狼蛛科各属间的遗传距离介于0.026￣0.200之间。2种建树方法均表明:科内的属及属内的种优先聚在一起;水狼蛛属相对马蛛属是狼蛛科中较为原始的类群,分化较早;獾蛛属作为1个单系群与熊蛛属合为1个并系,属于狼蛛亚科。狼蛛科6属间的分子系统关系为(Pirata(Hippasa(Trochsa Arctosa(Pardosa Wadicosa))))。     

基于16S rRNA部分序列中国蝙蝠科蝙蝠系统发生分析

蝙蝠科蝙蝠遍布全世界,是蝙蝠中种类最多的一个科.尽管从形态学、胚胎学和分子生物学等方面认为蝙蝠科内长翼蝠亚科应该提升到科、鼠耳蝠属应该提升到亚科的分类地位,但是其科内的系统关系长期以来一直处于争议之中.本文对蝙蝠科11种38个标本线粒体16S rRNA部分序列进行了测序,并结合以前报道的13种(属于7科13属)蝙蝠的线粒体16S rRNA部分序列构建了系统树,结果表明:长翼蝠亚科可以提升到科的分类地位、鼠耳蝠属提升到亚科的分类地位,这与前人报道的结果一致;相对于由鼠耳蝠亚科、彩蝠亚科和管鼻蝠亚科构成的分支,蝙蝠亚科和Antrozoinae是一个并系群.蝙蝠亚科内的亲缘关系也进行了进一步的讨论.     

The phylogenetic position of Dictyoglomus thermophilum based on 16S rRNA sequence analysis

Abstract The phylogenetic position of Dictyoglomus thermophilum has been determined by comparative sequence analysis of in vitro amplified 16S rRNA genes from the type strain as well as from a Dictyoglomus isolate. Results indicate that it forms a deep branch within the phylum of Thermotogales or may even represent its own phylum. It does not contain signature sequences within the 16S rRNA which could relate it to the Thermotogales group.     

Bacterial phylogeny based on 16S and 23S rRNA sequence analysis

Abstract: Molecular phylogeny increasingly supports the understanding of organismal relationships and provides the basis for the classification of microorganisms according to their natural affiliations. Comparative sequence analysis of ribosomal RNAs or the corresponding genes currently is the most widely used approach for the reconstruction of microbial phylogeny. The highly and less conserved primary and higher order structure elements of rRNAs document the history of microbial evolution and are informative for definite phylogenetic levels. An optimal alignment of the primary structures and a careful data selection are prerequisites for reliable phylogenetic conclusions. rRNA based phylogenetic trees can be reconstructed and the significance of their topologies evaluated by applying distance, maximum parsimony and maximum likelihood methods of phylogeny inference in comparison, and by fortuitous or directed resampling of the data set. Phylogenetic trees based on almost equivalent data sets of bacterial 23S and 16S rRNAs are in good agreement and their overall topologies are supported by alternative phylogenetic markers such as elongation factors and ATPase subunits. Besides their phylogenetic information content, the differently conserved primary structure regions of rRNAs provide target sites for specific hybridization probes which have been proven to be powerful tools for the identification of microbes on the basis of their phylogenetic relationships.     

基于12S,16S和28S rRNA基因序列的中国小毛瓢虫属系统发育分析

【目的】小毛瓢虫属Scymnus Kugelann昆虫主要捕食蚜虫、蚧虫等害虫,是一类经济上重要的天敌昆虫。目前针对小毛瓢虫属的系统发育研究尚属空白,亚属之间的系统演化关系尚不明确,为了建立合理的分类系统,亟需对小毛瓢虫属的亲缘关系进行研究和探讨。【方法】以华南农业大学馆藏的小毛瓢虫属5亚属共44种为研究对象,采用PCR技术对12S, 16S和28S rRNA基因的部分序列进行扩增;运用MEGA 7.0分析了小毛瓢虫属内12S, 16S和28S rRNA基因的碱基组成,基于K2P模型计算了小毛瓢虫属44种的种间遗传距离;采用最大似然法(maximum-likelihood, ML)和贝叶斯推断法(Bayesian-inference, BI)构建该属的系统发育树。【结果】扩增获得小毛瓢虫属44种的12S rRNA基因序列平均长度为356 bp, 16S rRNA基因序列平均长度为351 bp, 28S rRNA基因序列平均长度为315 bp;序列分析表明,12S rRNA基因的A, T, G和C平均含量分别为38.8%, 43.5%, 11.9%和5.8%, 16S rRNA基因的A, T, G和C平均含量分别为37.6%, 40.3%, 14.4%和7.7%, 28S rRNA基因的A, T, G和C平均含量分别为26.7%, 18.3%, 31.4%和23.5%;基于联合序列分析的种间遗传距离为0.004～0.276,平均遗传距离为0.115。系统发育分析结果表明,小毛瓢虫属为单系起源,而小毛瓢虫亚属Scymnus(Scymnus) Kugelann、毛瓢虫亚属Scymnus(Neopullus) Sasaji、小瓢虫亚属Scymnus(Pullus) Mulsant和拟小瓢虫亚属Scymnus(Parapullus) Yang均为并系起源。【结论】基于12S, 16S和28S rRNA基因序列的小毛瓢虫属系统发育分析显示传统的形态学分类体系与基于分子数据分析的结果部分不一致,这表明应该对该属内各亚属的鉴别特征进行全面检视,筛选并确立各亚属的形态指标,同时也表明该属内的亚属分类单元需重新厘定。     

16SrRNA二级结构可变区图形分析在放线菌分类中的应用*

采用16SrRNA可变区二级结构图形分析,比较了姜氏菌属及几个相关属种可变区二级结构的变化。结果表明,在9个可变区二级结构中茎的长度、环的数目和类型、茎的碱基对、以及环内部碱基均有不同。尤其在V5和V6两个区,这种差别尤为明显。这为姜氏菌属的建立提供了又一个证据,并认为16SrRNA可变区二级结构分析,可以应用于属以上原核生物的分类。     

Phylogenetic analysis of the family Rhizobiaceae and related bacteria by sequencing of 16S rRNA gene using PCR and DNA sequencer

Abstract The 16S rRNA gene sequences of 19 strains covering 97% of the molecules were determined for the members of the family Rhizobiaceae and related bacteria by PCR and DNA sequencer. The three biovars of Agrobacterium were located separately, whereas Agrobacterium rubi clustered with A. tumefaciens . Phylogenetic locations for the species of the genera Rhizobium, Sinorhizobium, Agrobacterium, Phylobacterium, Mycoplana (M. dimorpha), Ochrobactrum, Brucella and Rochalimaea (a rickettsia) were intermingled with each other with the similarity values higher than 92%. The family Rhizobiaceae should be redefined including the above-mentioned genera despite the ability for plant association and nitrogen fixation. Bradyrhizobium japonicum and Mycoplana bullata were far remote from the other species and should be excluded from this family.     

利用16S rRNA基因同源性分析鉴定两株明串珠菌

从酸马奶中分离出2株明串珠菌KLDS 5.0301和KLDS 5.0302,对2株菌的16S rRNA基因经PCR扩增测序,将测序结果同该属内菌株的16S rRNA序列作多序列比较,并建立明串珠菌属的系统发育树.结果表明,KLDS 5.0301的16S rRNA序列同L. garlicum的同源性百分比为100%.KLDS 5.0302的16S rRNA序列同L.mesenteroides LM2菌株的16S rRNA序列的同源性百分比为99.9%.根据系统发育树的结果,将KLDS5.0301鉴定为L.garlicum,KLDS 5.0302鉴定为L.mesenteroides.菌株KLDS 5.0301和KLDS 5.0302的16SrRNA序列已经在GeneBank申请国际序列注册号,分别为DQ239691和DQ297412.     

The phylogenetic position of Hydrogenobacter acidophilus based on 16S rRNA sequence analysis

Abstract Hydrogenobacter acidophilus strain 3H-1 is a thermoacidophilic, obligately chemolithoautotrophic, hydrogen-oxidizer isolated from a Japanese solfataric field. Strain 3H-1 requires elemental sulfur for growth. We used PCR to amplify the 16S rRNA gene of strain 3H-1, and sequenced the amplification product directly. Phylogenetic analyses show strain 3H-1 is closely related to Aquifex pyrophilus and may be located in the deepest branch within the eubacterial phylogenetic tree. Sulfur-dependency of the ancestral eubacterium is also discussed.     

Nuclease S1 analysis of eubacterial 5S rRNA secondary structure

Summary Single-strand-specific nuclease S₁ was employed as a structural probe to confirm locations of unpaired nucleotide bases in 5S rRNAs purified from prokaryotic species of rRNA superfamily I. Limited nuclease S₁ digests of 3- and 5-end-labeled [³²P]5S rRNAs were electrophoresed in parallel with reference endoribonuclease digests on thin allel with reference endoribonuclease digests on thin sequencing gels. Nuclease S₁ primary hydrolysis patterns were comparable for 5S rRNAs prepared from all 11 species examined in this study. The locations of base-paired regions determined by enzymatic analysis corroborate the general features of the proposed universal five-helix model for prokaryotic 5S rRNA, although the results of this study suggest a significant difference between prokaryotic and eukaryotic 5S rRNAs in the evolution of helix IV. Furthermore, the extent of base-pairing predicted by helix IV needs to be reevaluated for eubacterial species. Clipping patterns in helices II and IV appear to be consistent with a secondary structural model that undergoes a conformational rearrangement between two (or more) structures. Primary clipping patterns in the helix II region, obtained by S₁ analysis, may provide useful information concerning the tertiary structure of the 5S rRNA molecule.     

基于12S和16S rRNA序列的湍蛙属部分物种的系统发育关系

测定了湍蛙属 6个种共 10个种群 ,以及 4个外群种的线粒体 12S和 16SrRNA基因片段 ,比对后有94 0bp序列 ,发现 35 2个变异位点、 186个简约性位点。运用NJ法、MP法、ML法构建了系统关系树 ,各系统树一致表明内群为一单系群 ,分为两组 :第一组中 ,四川湍蛙两种群先聚合 ,再和棕点湍蛙聚为一支 ;第二组中 ,香港湍蛙和戴云湍蛙聚为一支 ,而香港大屿山离岛湍蛙种群首先与华南湍蛙相聚 ,再与武夷湍蛙构成姐妹支。研究结果表明 :香港地区增加 1种湍蛙分布 ;戴云湍蛙是一有效种 ;四川湍蛙的石棉和洪雅种群间遗传差异达到或超过其他种间的分歧水平。     

嗜盐菌HBCC-2的16S rRNA基因测序分析及其培养特性

从连云港台南盐场海盐生产区中分离纯化到一株嗜盐古菌HBCC-2,该菌株经PCR扩增后,测定其16S rRNA基因序列,采用BLAST软件对基因库中基因序列进行同源性比较,选取其相似性序列,采用Clustalx1.8和MEGA3.1软件对其16S rDNA序列进行了系统发育分析研究,结果表明HBCC-2菌株与菌株Halorubrum sp.GSL5.48的相似性达99%,结合其形态观察及生理生化反应特性,初步确定该菌株属于嗜盐红菌属(Halorubrum),菌株HBCC-2的16S rDNA序列已登陆到GenBank,其序列号为EF687739.通过比较不同NaCl浓度、pH和培养温度对该菌株生长的影响情况,研究了该菌株的生长特性,结果表明NaCl浓度为4mol/L、温度为35℃和pH为7.0的培养条件下其生长最佳.     

基于18S rRNA和线粒体16S rRNA基因序列的柳蚕进化分析

为探讨柳蚕Actias selene Hübner与鳞翅目昆虫的系统发育关系,本研究利用PCR扩增获得了柳蚕核糖体18S rRNA和线粒体16S rRNA基因的部分序列,长度分别为391bp和428bp。并采用邻近距离法(NJ)、最大简约法(MP)、类平均聚类法(UPGMA)构建系统进化树。结果表明,柳蚕线粒体16SrRNA基因序列与大蚕蛾科昆虫的16SrRNA基因序列均表现出偏好于碱基AT的倾向。柳蚕与所研究的其它蚕类的遗传距离介于0.016至0.140之间,其中与温带柞蚕Antheraea roylii的遗传距离最小,与野桑蚕Bombyx mandarina的遗传距离最大。而基于鳞翅目昆虫18S rRNA基因部分序列的进化分析显示,柳蚕与柞蚕Antheraea pernyi之间的遗传距离最小(0.010),与蓖麻蚕Samia ricini的遗传距离最大(0.017)。     

双歧杆菌地高辛标记寡核苷酸基因探针制备和应用研究

目的探讨地高辛标记寡核苷酸基因探针应用于微生态研究的可行性和实用性。方法制备双歧杆菌属和部分种的地高辛标记16S rRNA寡核苷酸探针,初步应用于微生态制剂鉴定和临床肠道微生态检测,评价寡核苷酸探针杂交在肠道微生态研究和检测中的应用价值。结果地高辛标记寡核苷酸探针具有较好的特异性与灵敏度：地高辛标记的双歧杆菌属和种的共6种寡核苷酸基因探针与标准菌株杂交后灵敏度和特异度分别为属探针95％、75％,青春双歧87．5％、90％,两歧双歧87．5％、87．5％,短双歧87．5％、92．5％,婴儿双歧75％、95％,长双歧75％、100％。结论寡核苷酸基因探针用于肠道细菌的鉴定显示出一定前景,加大探针的种类与扩大调查范围有可能使该技术替代现有细菌培养技术。     

16S rRNA analysis of Listeria monocytogenes and Brochothrix thermosphacta

Abstract Listeria monocytogenes and Brochothrix thermosphacta were investigated by the 16S rRNA cataloguing approach in order to determine their phylogenetic relationship. Both species are specifically, although moderately related, forming one of several sublines within the Bacillus-Lactobacillus-Streptococcus cluster of the ' Clostridium ' subbranch of Gram-positive eubacteria.