中国沿海日本囊对虾线粒体16S rRNA基因序列变异及遗传分化

对中国东南沿海4个日本囊对虾地理群体16S rRNA基因片段进行了序列测定和分析.在获得的467bp 序列中,共检测到45个变异位点,多态位点比例为9.64%.具有10种单倍型,三亚群体单倍型多样性最高(0.900±0.161).各群体的核苷酸多样性(π)为0.0007～0.0082,其中三亚群体核苷酸多样性最为丰富,远高于湛江、北海和海口群体.群体间遗传分化指数(F_(st))为0.000～0.945.结合已报道的16S rRNA基因序列,构建了NJ分子系统树,中国海域日本囊对虾可分为3支.香港至台湾沿岸所有单倍型聚集为一分支,湛江、海口和北海群体内的所有单倍型聚集成另一分支,两分支分别属于已报道的变种Ⅰ和变种Ⅱ,但三亚群体内的4个单倍型单独聚集成一分支,与前两分支的核苷酸序列差异大(7.95%～8.35%),遗传分化明显,可能为一亲缘关系很远的新种(或变种).研究结果可为日本囊对虾的资源管理和遗传选育提供参考.     

基于ITS2和16S rRNA的西施舌群体遗传差异分析

目前,我国西施舌群体分子遗传差异研究结果存在争议。分析我国南北沿海（9个群体）、与广西北海毗邻的越南（1个群体）西施舌核DNA的内转录间隔区2（ITS2）和线粒体DNA的16S rRNA基因（16S）片段核苷酸序列及其二级结构,为解决争议问题提供分子生物学资料。扩增获得西施舌ITS2片段和16S序列,其长度分别为389-402 bp和306 bp,加之下载序列共147条;序列分析显示,74个ITS2序列共有17种基因型,73个16S序列有15种单倍型,其中,长乐（CL）群体独享9种ITS2基因型和5种16S单倍型,非长乐群体（nCL）多数为群体间交叉共享1种或几种基因（单倍）型;基因（单倍）型核苷酸变异位点占5.7%（ITS2）和11.8%（16S）;基于ITS2和16S的CL群体和nCL群体间的遗传距离与群体内遗传距离之比分别为2.42和11.08,nCL群体间的平均遗传距离均为0.007;二级结构显示CL群体ITS2的9种基因型和16S的5种单倍型均区别于nCL群体,nCL的ITS2和16S二级结构分别相似;ITS2和16S基因的系统发育分析显示,CL西施舌形成支持率很高（98,96）的单系支,而nCL群体则交叉聚为另一支（98,96）。研究结果揭示,福建西施舌是腔蛤蜊属（Coelomactra）的一个新种。     

基于线粒体COⅠ和16S rRNA基因序列比较分析东海带鱼群体遗传多样性

测定了东海带鱼(Trichiurus lepturus Linnaeus)三个群体(命名为A: 122°32′E 29°55′N、B: 123°30′E 26°75N; C: 124°24′E 27°26′N)72个个体的线粒体DNA16S rRNA和COⅠ基因序列,通过单基因序列和联合序列分析, 研究了东海带鱼群体的遗传变异情况。分别得到1130 bp的COⅠ基因序列片段和554 bp的16S rRNA序列片段, 其中COⅠ基因片段的T、C、A、G含量分别为29.0%、28.9%、24.4%和17.7%; 16S rRNA基因片段的T、C、A、G含量分别为22.7%、27.6%、28.0%和21.7%。基于线粒体16S rRNA、COⅠ和16S+COⅠ基因序列分析, 72个个体中分别确定43个, 8个和49个单倍型, 存在单倍型共享现象。群体的单倍型多样性指数为0.9766—0.9992, 显示了群体内的单倍型较为丰富。3个群体间各序列平均核苷酸差异数(K)在5.111—9.024和0.0045—0.0076, 核苷酸多样性指数(π)为0.0048—0.0084, 显示不同带鱼群体遗传多态性丰富。使用邻接法构建的分子进化树揭示同一群体内大部分个体聚在一起。分析结果表明, 群体A遗传背景比群体B、群体C较为丰富, 群体内部个体差异大于群体间差异, 群体间基因交流频率较高, 遗传分化不明显, 初步判定东海带鱼3个群体的遗传多样性偏低。     

鹅喉羚线粒体D-loop和Cytb序列遗传多样性分析

[目的]为了探讨新疆天山野生动物园17只鹅喉羚的遗传多样性、亲缘关系和群体来源。[方法]对这些鹅喉羚线粒体控制区(D-loop)和细胞色素b(Cytb)基因序列进行了PCR扩增和测序,分别计算了群体单倍型多样度(H)和核苷酸多样性(Pi)以及个体间的遗传距离,并构建了单倍型NJ(Neighbor-Joining)聚类图。[结果]D-loop区和Cytb分别获得了978bp和543bp的基因片段,D-loop区识别出11个单倍型,Cytb区识别出3个单倍型;D-loop和Cytb的单倍型多样度(H)和核苷酸多样性(Pi)分别为0.863、0.01028和0.330、0.00077;个体之间最大遗传距离为0.024,最小遗传距离为0.001;17只鹅喉羚分别聚为2个分支。[结论]该鹅喉羚群体D-loop区多态性较丰富,在野外可能来自两个不同的群体。     

四川裂腹鱼乌江种群mtDNA控制区序列的遗传多样性分析

采用PCR扩增和直接测序法分析了四川裂腹鱼乌江种群32个个体mtDNA控制区的序列差异和遗传多样性.在该种群mtDNA控制区长度为464 bp的同源序列中,共计有9个变异位点,占总位点数的1.94%.32个个体共定义了9种单倍型,单倍型间平均遗传距离(P)为0.0060,单倍型多样度(Hd)为0.768.核苷酸多样性(π)为0.00324,平均核苷酸差异数(K)为1.500.用单倍型间遗传距离构建的NJ分子系统树聚为两个分支.结果提示四川裂腹鱼乌江种群遗传多样性匮乏,保护四川裂腹鱼乌江种群刻不容缓.     

基于线粒体COⅠ基因序列的东南沿海可口革囊星虫遗传多样性分析

为了解我国东南沿海可口革囊星虫自然群体的遗传多样性及遗传结构, 以线粒体COⅠ基因为分子标记, 对浙江象山(XS)与温岭(WL)、福建宁德(ND)、广东湛江(ZJ)4个可口革囊星虫自然群体的80个样本的COⅠ基因片段进行PCR扩增、序列测定和分析。结果表明, 在815 bp长度的核苷酸片段中, A、T、C、G碱基的平均含量分别为29.8%、31.0%、22.6%和16.6%, A+T含量(60.8%)高于C+G含量(49.2%), 表现出较强的AT偏好性。共检测到29个核苷酸变异位点, 定义了29种单倍型, 总群体单倍型多样性指数(H_d)、核甘酸多样性指数(P_i)及平均核苷酸差异数(K)分别为0.932、0.0036及2.8902, 表现出高的H_d和低的P_i。单倍型邻接关系树的拓扑结构简单, 未呈现明显的地理谱系结构。群体内的遗传距离为0.0027—0.0040, 群体间的遗传距离为0.0032—0.0040。两两群体间的遗传分化系数(F_st)和分子方差分析(AMOVA)表明, 可口革囊星虫的遗传变异主要来自于群体内, 而群体间无显著分化。中性检验和核苷酸不配对分布结果揭示, 可口革囊星虫经历了群体历史扩张事件, 大致发生在4.6万年前的更新世晚期。     

假眼小绿叶蝉地理种群的线粒体DNA 16S rRNA基因序列分析及遗传分化研究

测定了12个不同地理种群的茶树害虫假眼小绿叶蝉Empoasca vitis(Gothe)和1个外群共103个个体的线粒体DNA 16S rRNA基因部分序列,比较其同源性,计算核苷酸组成,并构建单倍型进化关系图。结果表明:在获得的假眼小绿叶蝉536 bp的序列中A+T含量占76.8%,其中56个核苷酸位点存在变异;100个个体共检测出56种单倍型,单倍型多样性指数(H)为0.971,核苷酸多样性指数(Pi)0.006;12个种群间的平均基因流(Nm)为1.64。总群体的固定系数Fst为0.026;AMOVA分子方差分析结果表明假眼小绿叶蝉的遗传分化主要来自于种群内部(97.4%)。各地理种群的遗传距离与地理分布不具有直接对应的关系。根据构建的单倍型进化关系网,各地理种群的单倍型呈现一种混杂的分布格局,未显示出与地理分布的一致性。     

福建畲族群体17个Y-STR基因座单倍型及遗传关系

应用Y-filerTM试剂盒及基因分型技术,检测152份福建畲族无关男性个体17个Y-STR基因座的多态性分布,计算等位基因频率及单倍型多样性,并结合已公开发表的其他11个群体相应基因座的单倍型资料,分析福建畲族群体遗传距离和聚类关系。福建畲族DYS385a/b基因座检出50种单倍型,其余15个Y-STR基因座分别检出3-11个等位基因,基因多样性GD值在0.4037(DYS391)～0.9725(DYS385a/b);观察到DYS19和DYS390基因座双等位基因和DYS385a/b基因座三等位基因,以及DYS448等部分基因座出现的"off-ladder"等位基因现象。17个Y-STR基因座共同构成的单倍型144种,其中138种单倍型出现1次,5种出现2次,1种出现4次,累计GD值为0.9990。从遗传距离分析发现,福建畲族与浙江汉族之间的遗传距离最近(0.0042),与青海藏族(0.2378)之间的遗传距离相对较远。福建畲族最靠近由台湾群体、浙江汉族、南方汉族等典型南方汉族群体聚成的分支区域。结果表明该17个Y-STR基因座在福建畲族群体中具有丰富的遗传多态性,对建立Y染色体STR数据库,研究群体遗传学和进行法医学应用有重要意义。     

南海野生斑节对虾的线粒体16S rRNA基因序列单倍型分析

采用聚合酶链式反应(PCR)技术对我国南海海域5个地点(三亚、深圳、阳江、湛江、北海)的野生斑节对虾100个个体的mtDNA 16S rRNA序列进行扩增,扩增产物经纯化后进行测序.用Clustal_X排序软件对所得的100个mtDNA 16S rRNA序列进行比对.通过ARLEQUIN软件对所得100个mtDNA 16S rRNA序列进行比较分析,共检测出28个变异位点,19种单倍型.19单倍型序列的碱基组成显示出较高的A T比例(69.0%).19种单倍型序列的遗传距离在0.002～0.033.根据构建的单倍型进化关系网,5个地点群体中的单倍型呈现一种混杂的分布格局.此外,根据单倍型在5个地点群体出现的频率和单倍型进化关系网,单倍型7是最为原始的单倍型,其他18种单倍型均起源于单倍型7.     

长江中下游褶纹冠蚌10个群体COI基因序列变异分析

以线粒体CO I基因为分子标记,对长江中下游褶纹冠蚌(Cristaria plicata)10个群体共200个个体的遗传多样性进行了研究.获得了620 bp的同源序列,A+T的平均含量为60.1%,明显高于G+C含量(39.9%),有105个核苷酸变异位点,占全部碱基数的17%,转换和颠换之比达到8.7,检测到了58个单倍型(GenBank登录号:EU698893～EU698950),Hap-5是主体单倍型,占总个体数的41%.褶纹冠蚌鄱阳湖群体(PY)平均核苷酸差异数和核苷酸多样性指数均最高,分别为25.426和0.041 01,太湖群体(TH)和衢州群体(QZ)遗传多样性参数较低.基于群体间遗传距离构建的NJ系统树中,洪泽湖内的3个群体与巢湖群体首先聚为一支,然后与由钱塘江群体、太湖群体、衙州群体和洪湖群体聚成的分支聚在一起,而后与洞庭湖群体聚在一起,最外侧一支为鄱阳湖群体.分子方差分析(AMOVA)得出群体间的遗传分化指数(Fst)为0.208 1(P<0.001),说明我国褶纹冠蚌不同群体间存在一定的遗传分化.     

Genetic diversity and population connectivity of the Asian green mussel Perna viridis in South China Sea,inferred from mitochondria DNA markers

The Asian green mussel Perna viridis is ecologically and economically important in the coastal regions of China. In order to characterize the genetic diversity and population connectivity of P. viridis in South China Sea, a 664 bp region of mitochondrial COI gene and a 293 bp region of 16S rRNA gene were sequenced and analyzed for 78 and 92 individuals from four populations in South China Sea, respectively. A total of 15 haplotypes were defined by 14 variable nucleotide sites in COI gene, and 7 haplotypes by 6 variable nucleotide sites in 16S rRNA gene. High haplotype diversity and low nucleotide diversity were observed in COI gene, while moderate haplotype diversity and low nucleotide diversity were observed in 16S rRNA gene. Pairwise F_ST values of COI gene were all negative and those of 16S rRNA gene ranged from −0.01409 to 0.10289. The results showed that no significant genetic divergence (or shallow genetic structure) and high levels of population connectivity among the four populations of P. viridis in South China Sea.     

Diversity of culturable actinobacteria isolated from marine sponge <Emphasis Type="Italic">Haliclona</Emphasis> sp.

This study describes actinobacteria isolated from the marine sponge Haliclona sp. collected in shallow water of the South China Sea. A total of 54 actinobacteria were isolated using media selective for actinobacteria. Species diversity and natural product diversity of isolates from marine sponge Haliclona sp. were analysed. Twenty-four isolates were selected on the basis of their morphology on different media and assigned to the phylum Actinobacteria by a combination of 16S rRNA gene based restriction enzymes digestion and 16S rRNA gene sequence analysis. The 16S rRNA genes of 24 isolates were digested by restriction enzymes TaqI and MspI and assigned to different groups according to their restriction enzyme pattern. The phylogenetic analysis based on 16S rRNA gene sequencing showed that the isolates belonged to the genera Streptomyces, Nocardiopsis, Micromonospora and Verrucosispora; one other isolate was recovered that does not belong to known genera based on its unique 16S rRNA gene sequence. To our knowledge, this is the first report of a bacterium classified as Verrucosispora sp. that has been isolated from a marine sponge. The majority of the strains tested belong to the genus Streptomyces and three isolates may be new species. All of the 24 isolates were screened for genes encoding polyketide synthases (PKS) and nonribosomal peptide synthetases (NRPS). PKS and NRPS sequences were detected in more than half of the isolates and the different "PKS-I-PKS-II-NRPS" combinations in different isolates belonging to the same species are indicators of their potential natural product diversity and divergent genetic evolution.     

Identification of Escherichia coli through analysis of 16S rRNA and 16S-23S rRNA internal transcribed spacer region sequences

A bacterial strain, designated BzDS03 was isolated from water sample, collected from Dal Lake Srinagar. The strain was characterized by using 16S ribosomal RNA gene and 16S-23S rRNA internal transcribed spacer region sequences. Phylogenetic analysis showed that 16S rRNA sequence of the isolate formed a monophyletic clade with genera Escherichia. The closest phylogenetic relative was Escherichia coli with 99% 16S rRNA gene sequence similarity. The result of Ribosomal database project's classifier tool revealed that the strain BzDS03 belongs to genera Escherichia.16S rRNA sequence of isolate was deposited in GenBank with accession number FJ961336. Further analysis of 16S-23S rRNA sequence of isolate confirms that the identified strain BzDS03 be assigned as the type strain of Escherichia coli with 98% 16S-23S rRNA sequence similarity. The GenBank accession number allotted for 16S-23S rRNA intergenic spacer sequence of isolate is FJ961337.     

Genetic analyses of Dinophysis spp. support kleptoplastidy

The question of whether the toxin-producing and bloom-forming dinoflagellate genus Dinophysis contains plastids that are permanent or contains temporary so-called kleptoplastids is still unresolved. We sequenced plastid 16S rRNA gene, the complete trnA gene and the intergenic transcribed spacer region located between the trnA gene and the 23S rRNA gene, and performed diagnostic PCR on cells of the genus Dinophysis. Dinophysis spp. were collected from five different geographical regions: the Baltic Sea, the North Sea, the Greenland Sea and the Norwegian fjord Masfjorden. In most cases the sequence analysis showed that the sequences were identical to each other and to sequences from the cryptophyte Teleaulax amphioxeia SCCAP K0434, regardless of the place of sampling or the species analyzed. The exception was some cells of Dinophysis spp. from the Greenland Sea. These contained a 16S rRNA gene sequence that was more closely related to the cryptophyte Geminigera cryophila. The cells of Dinophysis contained either one of the 16S rRNA gene sequences or both in the same cell. Our results challenge the hypothesis that the plastids in Dinophysis are permanent and suggest that they are more likely to be kleptoplastids.     

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。     

Genetic characterization of brown meagre (Sciaena umbra) and Shi Drum (Umbrina cirrosa) populations

This study analyzed population structure of fish, brown meagre (Sciaena umbra) and Shi drum (Umbrina cirrosa) from the Black Sea, the Aegean Sea, and the Mediterranean Sea. The Cytb and 16S rRNA genes of Shi drum and brown meagre fish were sequenced. In brown meagre and Shi drum, 25 and 20 haplotypes, respectively, of Cytb gene were identified; while for 16S rRNA, 4 and 8 haplotypes were identified. Nucleotide diversities of 16S rRNA and Cytb gene sites were found to be 83.8% and 52.6%, respectively, for brown meagre; while that for Shi drum were of 80.5% and 73.6%, respectively. There was a significant relationship between geographic distance and the genetic distance of the fish. Since Shi drum and brown meagre are migratory species, they can migrate between the seas. The lack of barriers among different populations facilitates the gene flow among the populations belonging to different regions. Since there is no information available on Shi drum and brown meagre population genetics, this study may be useful to understand the genetic diversity of these species to assist fishery managers for the management of these resources in terms of conservation and sustainability.     

新疆艾比湖嗜盐菌的细菌视紫红质和16S rRNA基因序列的研究

为了研究分析嗜盐古生菌物种与细菌视紫红质(BR)蛋白基因资源,从40份土壤、湖水及淤泥样品中分离出148株嗜盐菌,对其中6株菌采用聚合酶链式反应(PCR)方法对其编码螺旋C至螺旋G的蛋白基因片段和16SrRNA基因进行了扩增,并测定了基因的核苷酸序列。与已报道的相应片段进行对比,ABDH10,ABDH1I和ABDH40中的螺旋C至螺旋G的蛋白与其他菌株差异显著。基于16SrRNA序列的同源性比较以及系统发育学研究表明,ABDH10和ABDH40是Natronorubrum属下的新成员和Natrinema属下的新成员,ABDH40的16SrRNA序列已登录到GenBank,其序列号为AY989910。ABDH11中的螺旋C至螺旋G的蛋白与其他菌株差异显著。     

利用线粒体16S rRNA基因全序列分析直翅目主要类群的系统发生关系

为了构建稳健的直翅目主要类群间的系统发生关系并探讨16S rRNA基因序列在构建直翅目昆虫不同分类阶元系统发生关系时的可行性、功效以及性能,文章测定了直翅目4总科9科18种昆虫的16S rRNA基因全序列,联合已知该基因全序列的其他40种昆虫,构建了直翅目主要类群之间的系统发生关系,并分析了16SrRNA基因全序列的系统发生性能和功效。结果表明,直翅目昆虫的16S rRNA基因全长平均为1 310 bp;除生活方式特化的蚤蝼总科和蝼蛄总科的地位无法确定外,直翅目其他主要类群系统发生关系比较稳定;蝗总科下除了斑翅蝗科和槌角蝗科外,剑角蝗科、斑腿蝗科、网翅蝗科都不是单系群,且用不同的方法构建的系统发生树中聚类情况完全一致,各科间遗传距离差异不大,建议将其合为一科;锥头蝗科、瘤锥蝗科和癞蝗科间的遗传距离差异也不大;在构建系统发生树时,16S rRNA基因环区的信息量要比茎区的大;16S rRNA基因可以构建可靠的直翅目属与种水平和目与亚目高级阶元的系统发生关系,但对科和总科阶元缺乏足够的分辨力。     

甘蔗茎内两个内生菌株16S rRNA 序列分析及其生长特性

从两个甘蔗品种GT11和RB86-7515表面灭菌的茎中分离到两株具有固氮活性的菌株,分别编号为B11S和B8S.利用16S rRNA序列分析对其进行鉴定,并对两个菌株的生物学特性进行了比较.结果表明:B11S菌株与Stenotrophomonas maltophili菌株处在同一个分支,其序列与多个Stenotrophomonas maltophili菌株的序列相似性都达到98%以上,菌株B8S与多个土壤杆菌属细菌的序列相似性达到100%;两个菌株在温度31℃、pH为6左右时生长量最高;温度31℃、pH为6.5～7.0时固氮酶活性最高;相同浓度的蔗糖生长量大于葡萄糖,且表达固氮酶活性也最强;添加一定量的氮素有利于细菌生长和固氮酶活性的表达,但随着N含量的增加,抑制作用越来越明显.