木根麦冬与林生麦冬5S rRNA基因的进化

为了揭示多拷贝基因的进化方式,对濒危植物木根麦冬（Ophiopogon xylorrhizus Wang et Dai)3个居群、6个个体的294个5S rRNA基因拷贝及其姐妹种林生麦冬（O.sylvicola Wang et Tang) 的45个拷贝进行了DNA测序和序列分析,并以这个迄今发表的最大的单个物种的5S rDNA基因数据,以PAUP程序重建了分子系统发育树。结果表明：1）所得序列呈高度多样性,长度变化在307－548碱基之间,仅13对（3．8％）相同,序列分化指数较高：木根麦冬是0．078,林生麦冬是0．032,两物种间是0．149;2）100％的统计值支持两物种的5S rRNA基因分别来自于祖先种的一个拷贝,即“建立者拷贝”,这个拷贝在物种形成之后进行了一系列连续的扩增,形成一个直系的基因家族,而祖先种的其他拷贝在物种形成后被丢失;3）不同拷贝是独立进化的,序列间的一致化过程很弱,这在串联重复的rRNA基因中是罕见的;4）木根麦冬居群间曾存在频繁的基因交流,使5S rRNA基因的许多拷贝扩散于不同居群,维持着种内较高的遗传多样性。可以认为是某些近期发生的变化,阻止了居群间的基因交流,导致该物种广泛的自交,发生自交衰退,并最后导致濒危。     

不同地区麦冬遗传多样性的ISSR分析

利用ISSR分子标记技术对5个地区6个麦冬居群进行遗传多样性分析。选用10条扩增带型清晰且重复性好的引物进行扩增,共获得115条带,其中89条具有多态性,多态性比例为77.39%。当遗传相似系数(GS)为0.61时,可将6个麦冬居群分为2大类群。居群的GS平均值为0.643,表明供试居群之间存在较近的亲缘关系,且地域越近的其亲缘关系关系越近。研究表明ISSR分子标记技术能够很好地用于不同地域同种物种的亲缘关系分析。     

濒危植物木根麦冬的胚胎学研究

木根麦冬的花药壁分为4层,即:表皮、药室内壁、中层与绒毡层。绒毡层为腺质型。大孢子母细 胞减数分裂过程中的胞质分裂为连续型。小孢子四分体排列为左右对称型与四面体型。雌蕊子房壁在 授粉后4～5天破裂。胚珠倒生,双珠被,厚珠心。大孢子四分体线形排列,胚囊发育为蓼型。极核在受 精前融合成一个大的次生核。胚胎发育为紫菀型。核型胚乳。木根麦冬居群中存在雄性不育植株,约 占55．8％。胚胎在其发育早期约有43．3％败育。根据本研究的实验观察,(1)雄性不育和自花授粉以 及低效率的传粉机制而导致的异花传粉的失败;(2)部分胚胎的败育;(3)子房壁破裂而导致胚珠裸露等是木根麦冬有性生殖过程中的薄弱环节,也可能是导致该植物濒危的部分因素。     

木根麦冬（Ophiopogon xylorrhizus）干叶提取DNA用于RAPD分析

木根麦冬（Ｏｐｈｉｏｐｏｇｏｎｘｙｌｏｒｒｈｉｚｕｓ）是我国珍稀濒危植物,分布仅限于云南西双版纳雨林,在植物系统学和保护生物学研究中具有独特的意义。随机扩增多态ＤＮＡ（ＲＡＰＤ）方法是揭示群体遗传多样性的高效、简便方法,但一般均以新鲜材料提取总ＤＮＡ,对一些分布边远地区物种难以采用此法。本文研究从木根麦冬干叶片中提取总ＤＮＡ,进行ＲＡＰＤ分析。样品取自４个居群、４９个个体。选取生长旺盛的叶片,在野外用硅胶快速干燥保存样品。采用高盐低ｐＨ值法提取总ＤＮＡ,每克鲜重所得的干叶可得８０～１６０μｇ。通过对模板ＤＮＡ的各种处理和ＰＣＲ扩增程序的调整,解决了扩增片段边缘弥散、界线模糊、产率低等问题,获得了理想的扩增带型。这一成果对其它从野外直接采样的干叶提取ＤＮＡ进行ＲＡＰＤ研究具有指导意义     

青藏高原濒危植物唐古特大黄的遗传多样性

唐古特大黄(Rheum tanguticum)是中国传统的中藏药材,近几年由于生境的严重破坏,已濒临灭绝,并被列入濒危植物名单。为了探索唐古特大黄物种濒危的原因并保护其野生资源,本研究采集了9个居群87个个体的唐古特大黄样本,基于该物种的叶绿体基因trn S-G序列对其进行了遗传多样性研究。结果表明,唐古特大黄物种具有较高的遗传多样性水平(Ht=0.694),其中95.97%的遗传分化来自于居群间(G_(ST)=0.960),4.03%的遗传分化来自于居群内(Hs=0.028)。AMOVA分析也显示唐古特大黄居群间基因流较小(N_m=0.01),存在较高的遗传分化(F_(ST)=0.9631)。唐古特大黄较高的遗传多样性水平可能与该物种较长的进化史和生活史有关,居群间较高的遗传分化可能与高山地区特殊的地理环境和人类活动有关。根据研究结果,建议对唐古特大黄所有野生居群进行就地保护,同时收集种质资源开展异地繁殖工作,以保护物种的遗传多样性,维持其进化潜力。     

濒危植物版纳青梅保护遗传学研究初报

运用 2 0个 10碱基随机引物 ,对中国龙脑香科 (Dipterocarpaceae)特有的珍稀濒危植物版纳青梅 (VaticaguangxiensisX .L .Mo)进行了RAPD多态性分析。 3个自然居群和 1个迁地保护居群 (分布于云南和广西 )共扩增出2 31个位点 ,多态位点所占比例 (PPB)为 5 3.6 8% ;观察等位基因数na =1.5 36 8,有效等位基因数ne =1.2 878,Nei基因多样性指数h为 0 .16 86 ,居群内的遗传多样性水平较低。基于AMOVA和POPGENE的结果均表明居群内的遗传变异大于居群间的遗传变异。居群内的遗传变异为 5 5 .0 9% ,居群间的变异为 44 .91% (AMOVA) ;基因分化系数Gst为 0 .3746 (POPGENE) ,表明居群间存在高水平的遗传分化。研究结果对该濒危植物的保护有重要意义。考虑到低水平的遗传多样性和高水平的居群分化 ,通过居群间种子和幼苗的交换来促进基因流是可行的保护方案。迁地保护居群 (ML)不具最高的遗传多样性 ,表明为了保护此濒危物种的全部遗传变异 ,需要进一步采集更多个体补充到迁地保护居群中     

白花刺续断野生居群的叶绿体全基因组特征解析

白花刺续断在中国西藏是一种常用的药用植物,但其叶绿体全基因组的相关研究较少。为揭示该物种叶绿体全基因组的基本特征并探讨其谱系遗传结构,该研究利用Illumina测序平台对来自5个野生居群的10个白花刺续断个体进行二代测序,经组装、注释,得到10条完整的叶绿体全基因组序列,并对它们的基因组特征和居群间的谱系进化关系进行了初步研究。结果表明：(1)白花刺续断的叶绿体全基因组大小为155 335～156 266 bp,共注释113个基因,包括72个蛋白编码基因、30个tRNA基因和4个rRNA基因,其叶绿体基因组的大小、结构、GC含量及基因组成等方面在种内高度保守。(2)基因组比较分析表明,白花刺续断变异较大的片段均位于单拷贝区,且IR边界未出现明显的扩张和收缩。(3)群体遗传分析发现,白花刺续断的野生居群具有明显的地理遗传结构,不同居群间在遗传距离与地理距离上具有一定的相关性。研究认为,白花刺续断叶绿体基因组在种内居群水平上比较保守,且叶绿体基因组可在居群水平上揭示物种的地理遗传结构。这为后续开展刺续断属物种群体遗传学和系统发育基因组学研究奠定了基础。     

兰花蕉的遗传多样性研究

采用简单重复序列区间(ISSR,Inter-Simple Sequence Repeat)分子标记技术,对采自广东省的濒危植物兰花蕉(Orchidantha chinensisT.L.Wu)的7个居群137个个体进行遗传变异分析。用10个ISSR引物共扩增出清晰谱带101条,其中58条具有多态性,总多态位点百分率为57.43%。居群水平相对较低,多态位点百分率在6.93%-35.64%之间,平均为18.24%。经POPGENE1.31数据处理,结果表明:在物种水平上Nei基因多样性为0.1254±0.1686;Shannon信息指数为0.2000±0.2429;Nei基因分化系数为0.5481,表明54.81%的遗传变异分布在居群间,45.19%的遗传变异分布在居群内。物种居群间的遗传一致度在0.8855-0.9511之间。我们认为红花潭是其最适合生境,建议在此建立自然保护区;鉴于兰花蕉居群间出现了一定程度的分化,为最大限度地保护兰花蕉的遗传多样性,建议在自然居群间进行相互移栽,以提高群体间的基因交流。     

基于生物学物种定义探讨物种形成理论与验证的研究进展

物种形成是进化生物学研究的一个永恒主题, 由于生物群体进化是连续和动态的, 物种界限变得难于界定。本文首先讨论了3种地理物种形成模式(同域、邻域及异域), 并分析了近期报道的研究证据。其次, 评述了合子后生殖隔离机制的分子遗传基础和应用群体基因组数据分析的证据, 包括BDMI模型(Bateson-Dobzhansky-Muller incompatibility)、QTLs (quantitative trait loci)、霍尔丹法则及大X染色体效应。最后, 探讨了交配系统作为合子前隔离机制之一与物种形成的关系, 认为近交或自交通过扩大种群遗传结构分化, 增强不同交配系统的种群间不对称基因渐渗, 或种群间无基因渐渗等途径, 促进新物种形成。已知植物交配系统的演化更倾向于从异交(或自交不亲和)向自交(或近交亲和)方式, 花性状和基因组的分化推动形成所谓的自交综合征, 研究交配系统驱动或强化物种形成模式对认识植物物种形成机制有重要意义。     

杜氏盐藻泛素基因cDNA的克隆及系统进化分析

根据玉米,水稻等物种泛素序列设计一对简并引物.提取杜氏盐藻细胞的总RNA,利用RT-PCR方法扩增盐藻泛素基因的cDNA片断,回收两个长度不同的片断ubi-1和ubi-2,将其克隆到pMDl8-T载体上,测序后进行序列分析,为克隆杜氏盐藻泛素基因的cDNA序列并进行进化分析.结果 ubi-1经测序后得到一个完整拷贝(228 bp)和一个不完整的泛素cDNA序列(191 bp).Ubi-2经测序后得到两个拷贝(556 bp)和一个不完整的泛素cDNA序列(191 bp).盐藻3个不同拷贝泛素cDNA序列之间存在差异,但所编码氨基酸序列相同.盐藻泛素cDNA序列与其他物种的泛素cDNA序列具有高的同源(70%～85%),所推导的氨基酸序列与其他物种仅存在1～2个氨基酸的差异.进化分析显示,所分离的盐藻泛素基因与两个模式生物果蝇和衣藻的泛素基因共处一个进化支,彼此亲缘关系最近.盐藻泛素基因与其他物种的泛素基因可能来自共同的"祖先"基因.在进化中高度保守.     

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))))。     

Copy Number of Ribosomal Operons in Prokaryotes and Its Effect on Phylogenetic Analyses

Different aspects of the presence of multiple copies of ribosomal operons in prokaryotic genomes are reviewed. The structure of prokaryotic ribosomal operons is briefly described. The available data are summarized regarding the copy number of ribosomal genes in various prokaryotic genomes, the degree of polymorphism of their individual copies, and physiological and evolutional aspects of the presence of the multiple copies of ribosomal genes. The review also considers the influence of the presence of multiple copies of ribosomal genes on the results of identification of prokaryotic isolates and of the studies of prokaryotic diversity in environmental samples based on phylogenetic analysis of 16S rRNA gene sequences.     

Embryological Studies on Endangered Ophiopogon xylorrhizus (Liliaceae)

The embryological studies on an endangered species Ophiopogon xylorrhizus are reported in this paper. The anther was four-sporangiate, and its wall was composed of four layers: epidermis, fibrous endothelium, middle layer and glandular tapetum. Cytokinesis of microspore mother cell in meiosis was of the Successive type, and tetrads were tetrahedral or isobilateral. The mature pollen grains were 2-celled. Ovules were anatropous, bitegminous, and crassinucellate. Ovary wall ruptured 4～5 days after fertilization. The development of embryo sacs conformed to the Polygonum type. The two polar nuclei fused into a secondary nucleus before fertilization. The embryogeny was of the Asterad type, and endosperm development corresponded to the Nuclear type. About 55.8 % individuals in the population of O. xylorrhizus were found male sterile, and 43.3% of embryos aborted at early development stage. As a conclusion, we consider that (1)the failure of the xenogamy resulted from male sterility, autogamy and inefficient pollination agents, (2)embryo abortion and (3)exposure of young seeds during their early development are the weak links in the sexual reproductionof O. xylorrhizus, which would contribute to the endangerment of this species.     

从线粒体基因探讨中国大头蛙群的分类及其属内地位

Partial sequences of the mitochondrial 12S rRNA and 16S rRNA gene were determined for 8 populations of three species of Chinese Limnonectes, and aligned with the published sequences of Limnonectes from other parts of the world. When Nanorana parker, Paa boulengeri, Fejervarya limnocharis and Hoplobatrachus rugulosus was used as outgroup taxa (Accession Nos. AY158705, AY313685, AF206111, AF206491, AY322311). The sequences of the 12S rRNA and 16S rRNA genes totaled 950 nueleotide positions with gaps including 510 variable sites. We reconstructed phylogenetie trees using Clustal X 1.8, Mega 2.1 and PHYLIP 3.5e software, and using the maximum parsimony and maximum likelihood methods, respectively. Our analyses suggest that these fanged frogs from China are another monophyletie group in addition to the four monophyletie groups identified by previous studies. The Chinese Limnonectes were grouped into three elades (BCL 55% ). The first elade contains one species (BCL 100% ), from a population of Limnoneetes fragilis from Hainan Province. The second contains four individuals (BCL 100% ), i. e. two populations of Limnonectes kuhlii from Yunnan Province. The third contains one species (BCL 100% ), i. e. five populations of Limnonectes fujianensis from Fujian Province and 1 from Taiwan Province. The resulted phylogenetie trees indicate L. fragilis is basal to L. kuhlii L. fujianensis 。     

用12S rRNA基因序列研究斑腿蝗科二属六种的进化关系

采用DNA测序技术测定了中国斑腿蝗科昆虫6种和斑翅蝗科的红胫小车蝗线粒体12S rRNA基因长约345 bp片段的序列。在获得的345 bp的序列中,A+T约占71.8%,其中135个核苷酸位点存在变异 (约占39.1%)。PAUP4.0b数据分析软件构建该6种蝗虫的MP和NJ分子系统树显示,稻蝗属和蔗蝗属各为独立的一支。在稻蝗属一支中,中华稻蝗与山稻蝗关系很近,而与小稻蝗关系较远,这与形态学结果相吻合;在蔗蝗属一支中,异歧蔗蝗与斑角蔗蝗亲缘关系较近,而与等歧蔗蝗关系较远,这与形态学研究结果并不吻合,有待进一步的研究。     

裸子植物5S rRNA基因序列变异及二级结构特征

在高等植物中,5SrRNA基因一级结构是高度保守的,二级结构也相当一致。通过比较18种裸子植物5SrRNA基因序列和二级结构变异,发现55％的核苷酸位点是可变的,这种变异有68％发生在干区（双链区）,其中一些变异,如双链的互补性核苷酸替代,GU配对等能够维系5SrRNA二级结构的稳定性。环区相对保守,这与5SrRNA三级结构折叠或在转录翻译过程中蛋白质、RNA的结合相关。另外,首次报道了松属环E区核苷酸的变异性,这可能与其他区域的变异一样,是假基因造成的结果。5SrRNA基因信息可反映大分类群的系统进化关系,但由于基因长度短,信息量小,其在近缘种系统分类的应用受到限制。     

Molecular systematics of the South American caviomorph rodents: relationships among species and genera in the family Octodontidae

Nucleotide sequences from mitochondrial (12S rRNA) and nuclear (growth hormone receptor) genes were used to investigate phylogenetic relationships among South American hystricognath rodents of the superfamily Octodontoidea, with special emphasis on the family Octodontidae. Relationships among most taxa were well resolved by a combined analysis of both genes, and the molecular phylogeny was used to address several long-standing phylogenetic problems. The family Abrocomidae was the most basal lineage within the superfamily Octodontoidea, sensu stricto, and the family Ctenomyidae was sister to the family Octodontidae, followed by a monophyletic group containing the families Myocastoridae and Echimyidae. A basic dichotomy was observed within the family Octodontidae. The Argentine desert specialists, Tympanoctomys and Octomys, grouped separate from Octodontomys, which was sister to a clade containing a monophyletic Octodon and a clade represented by species of Aconaemys and Spalacopus. Aconaemys was paraphyletic relative to Spalacopus. The phylogeny was used as an interpretive framework for an examination of variation in several non-molecular characters. The primitive diploid number for most of the octodontoids was determined to be between 46 and 56, and the primitive genome size 8.2 pg. Members of the Octodontidae appeared to be derived from an ancestral stock occupying lower elevations in scrub habitat. Furthermore, estimates of divergence time from the molecular data provided a temporal perspective for changes in plant communities, which demonstrated turnover and diversification in response to climatic and geologic events occurring in the Miocene through the Pleistocene.     

Scorpions of the genus Odontobuthus Vachon, 1950 (Scorpiones: Buthidae) from Iran: Phylogenetic relationships inferred from mitochondrial DNA sequence data

The molecular phylogeny of the genus Odontobuthus Vachon, 1950 (Scorpiones: Buthidae) in Iran was evaluated using two mitochondrial DNA genes, cytochrome c oxidase, subunit I (COI) and 16S ribosomal RNA (16S rRNA). The molecular phylogenetic analyses were performed using Maximum Parsimony, Maximum Likelihood and Bayesian inference methods. The resulting topologies supported two main clades: the clade comprising Odontobuthus doriae, O. bidentatus, and O. tavighiae, and another one which is the O. tirgari clade. The results clearly presented additional support for the taxonomic validity of the recently described species, O. tirgari and O. tavighiae. In addition, the monophyly of two previously described species O. doriae and O. bidentatus was confirmed. According to the data presented here, three taxonomically valid species belonging to the genus Odontobuthus occur in Iran.