植物核基因组核糖体基因间隔区序列的结构特点及其在系统发育研究中的应用

目前,在植物系统发育研究中应用较多的核基因组的核糖体DNA基因间隔区序列主要有5S rDNA基因间隔区、内转录间隔区ITS和基因间间隔区IGS.虽然这些间隔区序列在长度、结构等方面各不相同,但都具有进化速率较快的特点,在植物属及属下分类水平的系统发育关系研究中非常有用.本文重点就核基因组的5S rDNA基因间隔区以及IGS在植物中的特点以及各自在植物系统发育研究中的应用进行了综述.     

山羊草属二倍体物种核rDNA ITS区序列及其系统发育关系分析

测定了山羊草属（Ａｅｇｉｌｏｐｓ）二倍物种核ｒＤＮＡ ＩＴＳ区序列,发现其碱基粉介于６０１－６０７之间,比报道的小ｒｉｕｃｅａｅ）其他属的ＩＴＳ区我略长,Ｇ＋Ｃ含量达６１．１％～６２．９％,序列间的分化距离为０．００５０～０．０４６８。用ＰＨＹＬＩＰ３．５ｅ软件包对所测得的ＤＮＡ数据进行聚类分析,结果显示：１．Ａｅ．ｓｐｅｌｔｏｉｄｅｓ与该组其他种相距很远,支持将其从Ｓｉｔｏｐｓｉｓ组中独立出来,     

基于rDNA ITS序列对绒泡菌目黏菌系统发育的探讨

绒泡菌目Physarida是黏菌纲Myxogastria最大的一个目,对其系统发育关系的研究一直是根据形态特征。为了从分子水平探讨绒泡菌目乃至黏菌纲的系统发育关系,以黏菌r DNA ITS通用引物对绒泡菌目5属8种黏菌的r DNA ITS进行扩增和测序,结合Gen Bank中已有的黏菌r DNA ITS序列,利用贝叶斯推断法(Bayesian inference,BI)和最大似然法(Maximum likelihood,ML)构建系统发育树。结果表明:绒泡菌目不同物种的r DNA ITS区在碱基组成和长度上差异明显,长度为777–1 445bp,G+C mol%在53.4%–61.9%之间。绒泡菌目与发网菌目Stemonitida聚类为两个明显的分支,在绒泡菌目分支上,绒泡菌科Physaraceae和钙皮菌科Didymiaceae各聚为一支,支持了形态学上以孢丝是否具有石灰质为依据区分这两个科的观点。由多份不同地理来源的鳞钙皮菌Didymium squamulosum材料组成的钙皮菌科又形成3个分支,证实了这个形态种是由地域来源广泛、繁殖亲和性各异和遗传变异较大的不同生物种组成的复合体。     

核rDNA的ITS序列在被子植物系统与进化研究中的应用

被子植物与其它高等真核生物相似,核rDNA是高度重复的串联序列。由于同步进化的力量,绝 大多数物种中这些重复单位间已发生纯合或接近纯合。核rDNA的内转录间隔区(ITS)包含被5.8S rDNA所分隔的ITS1和ITS2两个片段,ITSl的长度为187～298bp,ITS2为187～252bp,经PCR扩 增后可以方便地对这两个片段进行直接测序或克隆测序。ITS序列变异较快,可以提供较丰富的变异 位点和信息位点,已证实它是研究许多被子植物类群系统与进化的重要分子标记,不仅可用于解决科、 亚科、族、属、组内的系统发育和分类问题,而且可用于重建多倍体复合体的网状进化关系,探讨异源多倍体的起源过程,然而,正是由于ITS序列变异较快,它一般不适于科以上水平的系统发育研究。     

依据ITS rDNA序列探讨我国盾腹虫属种类的系统发育关系(英文)

盾腹吸虫为寄生扁形动物中一小的类群。我国已报道7种盾腹吸虫,其中5种隶属于盾腹虫属(Aspidogastridae,Aspidogastrinae)。研究测定了在我国采集到的4种盾腹虫属吸虫的核糖体DNA转录内间隔区(ITSrDNA)序列,并分别采用邻接法和最大似然法构建分子系统发育树。结果显示,这4种盾腹吸虫的ITS-1和ITS-2序列的长度分别在728—877bp和518—645bp之间,其G+C含量分别在50.1%—52.3%和49.2%—52.2%范围内。4种盾腹吸虫的种间遗传距离在0.2%—26.9%之间,其中重庆盾腹吸虫(Aspidogaster chongqingensis)和似螺盾腹吸虫(A.limacoides)间仅为0.2%。所构建的最大似然树和邻接树具有相同的拓扑结构,均支持重庆盾腹吸虫和似螺盾腹吸虫亲缘关系最近,它们与饭岛盾腹吸虫(A.ijimai)亲缘关系较近,而与位于系统树基部的贝居盾腹吸虫(A.conchicola)关系较远。此外,对我国盾腹属种类的宿主特异性进行了讨论。     

虎耳草属山羊臭组的界定和系统发育: 核糖体DNA ITS序列证据

虎耳草属Saxifraga山羊臭组sect. Ciliatae是该属中最大的一个组, 共有175种, 主要分布在喜马拉雅地区, 我国分布有166种, 占总种数的95%; 其中, 112种为中国特有。约80%的种类分布在我国青藏高原和西南地区, 是中国喜马拉雅植物成分的代表类群。山羊臭组内物种分化十分显著, 分类处理也很困难, 该组是否为单系类群, 组下的系统发育关系也不清楚, 均需进一步验证。本文测定了虎耳草属山羊臭组及其他组33种植物样品的核糖体DNA内转录间隔区ITS序列, 并从GenBank调取虎耳草组sect. Saxifraga等组和近缘属唢呐草属Mitella共22种植物的该序列。ITS分析结果表明: (1)所研究的山羊臭组类群聚为单独一支, 而且与垫状组sect. Porphyrion、虎耳草组、球茎组sect. Mesogyne和仅在欧洲分布的sect. Cymbalaria和sect. Cotylea等8个组聚成的另一分支构成姊妹群; (2)根据形态特征建立的山羊臭组的3个亚组即唐古拉亚组subsect. Hirculoideae、莲座状亚组subsect. Rosulares和具芽亚组subsect. Gemmiparae各自聚为一支, 但是莲座状亚组这一支的支持率较低。同时, 山羊臭组的鞭匐枝亚组subsect. Flagellares和subsect. Hemisphaericae的代表类群单独聚为一支, 位于具芽亚组类群分支内部而不能成立; (3)唐古拉亚组和莲座状亚组又聚为一亚分支与具芽亚组构成姊妹群, 而且具芽亚组最早从山羊臭组这一支中分化出来。我们的研究还发现山羊臭组内种间形态分化较大, 而ITS碱基变异较小, 这可能是山羊臭组类群在青藏高原及毗邻地区的高山环境下物种快速分化的结果。     

基于ITS序列的苎麻属大叶苎麻组的系统发育研究

大叶苎麻组(Sect.Duretia)是荨麻科苎麻属(Boehmeria)的一个种类比较多的组,目前对该组植物的系统发育还不清楚.本研究测定了其中9种4变种的核糖体DNA的ITS序列.ITS平均长度约687 bp.选取冷水花属作为外类群,根据ITS序列的差异计算出种间遗传距离.分别采用最简约法和邻接法进行系统发育分析.细野麻(Boehmeria gracilis)、赤麻(B.silvestrii)、野线麻(B.japonica)和疏毛水苎麻(B.pilosiuacula)、束序苎麻(B.siamensis)亲缘关系较近聚成一支,自展支持率为76%(MP)和89%(NJ);密球苎麻(B.densiglomerata)、长序苎麻(B.dolichostachya)、灰绿水苎麻(B.macrophylla var.canescens)、糙叶水苎麻(B.macrophylla var.scabrella)、圆叶水苎麻(B.macrophylla vat.rotundifolia)和海岛苎麻(B. formosana)、福州苎麻(B.formosana var.stricta)则聚为另一支,自展支持率为74%(MP)和94%(NJ);歧序苎麻(B.polystachya)单独为一支,与大叶苎麻组的其他种在一级分支中就分开,和其他种亲缘关系较远.     

黑粉菌属某些种的核rDNA ITS2序列分析

本文报道从某些黑粉菌的黑粉孢子粉末中提取核糖体脱氧核酸(rDNA)后,用聚合酶链式反应(PCR)的方法,对其转录间区(ITS2)片断进行了扩增,核苷酸序列测定和分析,其结果表明异形黑粉菌(Ustilago anomala)与其它3种近似黑粉菌,科尔达黑粉菌(U. cordae),柳叶刺黑粉菌(U. bungeana)和网孢黑粉菌(U. reticulata)的rDNA ITS2序列区别很大,在被测定的190个碱基对中,差别达69个位点以上(26.3-36.3%),异形黑粉菌与这三种黑粉菌亲缘关系较远,在中国作者原定名为异形黑粉菌的种与在欧洲取材的科尔达黑粉菌相似程度较高.因此将在中国已报道的异形黑粉菌更正为科尔达黑粉菌,而柳叶刺黑粉菌与科尔达黑粉菌近似程度也较高,将柳叶刺黑粉菌作为科尔达黑粉菌的异名.     

Species identity and phylogenetic relationship of the pearl oysters in Pinctada Röding, 1798 based on ITS sequence analysis

Analysis of ITS 1 and ITS 2 sequences in the pearl oysters Pinctada albina, Pinctada chemnitzi, Pinctada fucata, Pinctada fucata martensii, Pinctada imbricata, Pinctada margaritifera, Pinctada maxima, Pinctada nigra and Pinctada radiata was carried out. Homogeneity test of substitution patterns suggests that GC contents are highest in P. margaritifera and P. maxima and chromosomal rearrangements occurred in P. chemnitzi. These observations indicate that P. margaritifera and P. maxima are primitive species and P. chemnitzi is a recent species. Phylogenetic analysis shows that the pearl oysters studied constitute three clades with P. margaritifera and P. maxima forming the basal clade, congruent with results revealed by the substitution pattern test. The second clade consists of P. fucata, P. fucata martensii and P. imbricata. Low genetic distances among these taxa indicate that they may be conspecific. The remaining species make up the third clade and low genetic divergence between P. albina and P. nigra suggests that they may represent the same species. The ITS 1 sequence of P. radiata in GenBank is almost identical to that of P. chemnitzi determined in the present study and we suspect that the specimen used for the P. radiata sequence was misidentified.     

Molecular analyses of the IGS & ITS regions of rDNA of the psychrophilic yeasts in the genus Mrakia

Species of the genus Mrakia are currently classified as synonyms based on molecular sequence analyses of the large sub-unit ribosomal DNA (LrDNA). Physiological and protein electrophoretic studies, however, reveal possible species differences. To clarify this discrepancy, we undertook molecular sequence analyses of the internal transcribed spacer (ITS) and intergenic spacer (IGS) regions of rDNA from the four psychrophilic Mrakia species and the psychrophilic yeast, Cryptococcus curiosus. Identical ITS sequences were found between C. curiosus, M. nivalis and M. frigida. Although, M. stokesii and M. gelida displayed identical ITS and IGS sequences, their sequences differed from the other three species by 2.3% and 38%, respectively. The results suggest that M. stokesii is a synonym of M. gelida, whereas M. nivalis is a synonym of M. frigida. Sequence differences (1.9%) observed in the IGS region indicates that C. curiosus is a distinct strain of M. frigida.     

Phylogeny of slug species of the genus Arion: evidence of monophyly of Iberian endemics and of the existence of relict species in Pyrenean refuges

The Iberian Peninsula contains the majority of the Paleartic land slug species of the genus Arion, which exhibits diverse taxonomic problems. The present study investigated Arion taxonomy on the basis of analyses of the mitochondrial ND1 gene and nuclear internal transcribed spacer 1 (ITS1) sequences. The Iberian endemic species were monophyletically clustered in two divergent sister clades. The topotype specimens of Arion lusitanicus and the closely related species Arion nobrei and Arion fuligineus, as well as Arion hispanicus and Arion flagellus, were grouped into an ‘Atlantic’ clade, whereas Arion baeticus, Arion gilvus, Arion anguloi, Arion wiktori and Arion paularensis were included in a ‘Continental–Mediterranean’ clade. Calibration of mutation rate in the ND1 gene suggested that the divergence of these two clades occurred around the Pliocene–Pleistocene boundary, with subsequent speciation events during the Pleistocene. A group of ancestral and divergent endemic species with distribution centred in the Pyrenean mountain range (Arion molinae, Arion lizarrusti, Arion antrhacius and Arion iratii) arose in the Pliocene and survived through the Pleistocene in geographically confined small populations. Arion lusitanicus showed up to be polyphyletic: specimens, sampled outside the geographic range of the topotype in the north‐western Iberian Peninsula, were included in a non‐monophyletic clade together with the widely distributed species Arion ater and Arion rufus. The divergent species with a wide European distribution (Arion subfuscus, Arion hortensis, Arion fagophilus and Arion intermedius) were located in basal positions in all topologies. The evolutionary history of these slug species (highly sensitive to climatic factors, with capacity for both outcrossing and selfing, and with low dispersal ability) appears to have been moulded by Pliocene–Pleistocene climate events and by the rugged topography of southern Europe, giving rise to repeated cycles of population isolation during periods of glaciation alternating with interglacial expansions limited by geographic barriers.     

Geographical diversification of the genus Cicer (Leguminosae: Papilionoideae) inferred from molecular phylogenetic analyses of chloroplast and nuclear DNA sequences

Cicer L. (Leguminosae: Papilionoideae) consists of 42 species of herbaceous or semi-shrubby annuals and perennials distributed throughout the temperate zones of the Northern Hemisphere. The origin and geographical relationships of the genus are poorly understood. We studied the geographical diversification and phylogenetic relationships of Cicer using DNA sequence data sampled from two plastid regions, trnK / matK and trnS - trnG , and two nuclear regions, the internal transcribed spacer (ITS) and external transcribed spacer (ETS) regions of nuclear ribosomal DNA, from 30 species. The results from the phylogenetic analyses of combined nuclear and chloroplast sequence data revealed four well-supported geographical groups: a Middle Eastern group, a West-Central Asian group, an Aegean–Mediterranean group, and an African group. Age estimates for Cicer based on methods that do not assume a molecular clock (for example, penalized likelihood) demonstrate that the genus has a Mediterranean origin with considerable diversification in the Miocene/Pliocene epochs. Geological events, such as mountain orogenesis and environmental changes, are major factors for the dispersal of Cicer species. The early divergence of African species and their geographically distinct region in the genus suggest a broader distribution pattern of the genus in the past than at present.  © 2007 The Linnean Society of London, Botanical Journal of the Linnean Society, 2007, 154 , 175–186.     

Molecular phylogenetics of Hippophae L. (Elaeagnaceae) based on the internal transcribed spacer (ITS) sequences of nrDNA

 The genus Hippophae comprises 7 species and 8 subspecies according to the latest classification, and has shown enormous ecological, nutrient and medicinal values. Here we analyzed the phylogenetic relationships among 15 taxa of the genus by comparing sequences of the internal transcribed spacer (ITS) region of nuclear ribosomal DNA (nrDNA). ITS sequences in Hippophae varied in length from 651 bp to 666 bp. The aligned sequences were 690 bp in length and 269 (39.0%) were variable sites with 150 being parsimony-informative. The amount of polymorphism observed within a taxon was extremely low in most taxa except for two putative hybrid species. The aligned sequences were analyzed by maximum parsimony (MP) and neighbor-joining (NJ) methods. In the strict consensus trees of parsimony analysis, the monophyly of Hippophae was supported by 100% bootstrap value. H. tibetana was at the basal position of the genus, and the remaining taxa formed two clades with high bootstrap support. The first clade included subspecies of H.␣rhamnoides and the other one consisted of remaining species. Parsimony analysis also suggested that the species H. tibetana, H. neurocarpa and H.␣salicifolia were all distinct. Although the sequence divergence among subspecies of H. rhamnoides was also remarkably high, the molecular data supported the monophyly of H. rhamnoides when H. rhamnoides subsp. gyantsensis Rousi was excxluded. The NJ trees showed essentially the same topology. The taxonomical arrangement that divided the genus into two sections was not supported based on the ITS sequences. However, the hybrid origin of H. goniocarpa and H. litangensis proposed previously was supported by the present ITS data. Received January 7, 2002; accepted May 10, 2002 Published online: November 22, 2002 Addresses of the authors: Kun Sun, Xuelin Chen, Ruijun Ma, Qin Wang, Institute of Botany, Northwest Normal University, Lanzhou 730070, China. Changbao Li, Song Ge (e-mail: gesong@ns.ibcas.ac.cn or song_ge@hotmail.com), Laboratory of Systematic and Evolutionary Botany, Institute of Botany, Chinese Academy of Sciences, Beijing 100093, China.     

新疆石竹属野生种核糖体DNA的ITS序列与亲缘关系

新疆是我国石竹属植物分布和分化中心,种质资源丰富。通过采用PCR直接测序法,对新疆石竹属（Dianthus)植物野生种共8个种及外类群（Lychnis coronata Thunb)rDNA的ITS区（包括ITS－1,5.8S,rDNA和ITS－2）进行序列测定。研究结果说明,新疆石生属植物的ITS序列总长度为617－621bp,长度变异较小,仅相差4bp,种间序列同源性很高,达97．6％－99．8％,外类群的序列同源性为80％左右。ITS区序列在石竹属内是相当保守的,石竹属物种间序列变异位点基本上是转移多于颠换,且转移率较高,转换／颠换率为1．0－3．0。系统地位和亲缘关系分析表明分布于中国的石竹属植物3个组即齿瓣组（sect.Barbulatum Williams)和石竹组（sect.Dianthus),遂瓣组（sect.Fimbriatum Williams)的亲缘关系较远,而sect.Dianthus与sect.Fimbriatum Williams的亲缘关系较近。ITS系统发育树揭示了石竹组起源早于遂瓣组和齿瓣组,其结果与传统形态学论述存在很大差异。     

Tribal delimitation and phylogenetic relationships of Loteae and Coronilleae (Faboideae: Fabaceae) with special reference to Lotus: evidence from nuclear ribosomal ITS sequences

The temperate herbaceous tribes Loteae and Coronilleae have traditionally been regarded as taxonomically distinct entities. More recent morphological assessments, however, have challenged this view and suggest combining the two tribes under Loteae. Two key features used to distinguish the Coronilleae from Loteae include jointed fruits and branched root nodules. We evaluate the taxonomic utility of these characters using information derived from phylogenetic analyses of the internal transcribed spacers ITS1 + 2, and the intervening 5.8S region of nuclear ribosomal DNA. Results from this study show that neither the Loteae nor Coronilleae form individual monophyletic groups, and that key fruit and root nodule characters used to distinguish the Coronilleae are homoplastic. Given these data, we support the recognition of a single tribe, Loteae. We also find that Lotus, the largest and most morphologically complex genus in either tribe, is not monophyletic. Rather, it consists of two geographically distinct lineages, Old and New World, each of which are more closely related to other Loteae genera: Old World Lotus are more closely related to Old World Anthyllis, while New World Lotus show closer affinities to Old World Coronilla. These data also have important implications for the biogeography of New World Lotus: equally most parsimonious reconstructions suggest a complex scenario of intercontinental dispersals that involve not only Old World Lotus but Coronilla as well.     

Phylogeny of the Neotropical Alibertia group (Rubiaceae), with emphasis on the genus Alibertia, inferred from ITS and 5S ribosomal DNA sequences

Phylogenetic relationships of 38 species of the Alibertia group (Rubiaceae) and two outgroup species were investigated using the nuclear ribosomal 5S nontranscribed spacer (5S-NTS) and the internal transcribed spacers (ITS). Analysis of the data sets separately and in combination resulted in several well-supported and congruent groupings. However, the three analyses yielded different results as to the branching order of the basal clades. With the exception of Alibertia hispida, the species in the genus Alibertia appear in one weakly to moderately supported clade. This clade is in turn composed of two strongly supported subclades. One comprises several Alibertia species, including the type (A. edulis), three Borojoa species, and Randia tessmannii. The other subclade consists of Alibertia species only. This division is also generally supported morphologically by fruit size, corolla size, number of corolla lobes, and pollen aperture (porate vs. colporate). The sister group to the Alibertia clade comprises Duroia with Amaioua species internested. The close relationship of Ibetralia and Kutchubaea is corroborated. In addition, Alibertia hispida is a member of this strongly supported clade. Likewise, the two "Genipa" species are supported as a monophyletic group in 100% of the bootstrap replicates. It is concluded that the 5S spacer is superior to the commonly used ITS region in terms of resolution and robustness among closely related taxa.     

Review of the sawfly genus Empria (Hymenoptera, Tenthredinidae) in Japan

The following eleven Empria species are reported from Japan: Empria candidata (Fallén, 1808), Empria japonica Heidemaa & Prous, 2011, Empria liturata (Gmelin, 1790), Empria loktini Ermolenko, 1971, Empria plana (Jakowlew, 1891), Empria quadrimaculata Takeuchi, 1952, Empria rubicola Ermolenko, 1971, Empria tridens (Konow, 1896), Empria tridentis Lee & Ryu, 1996, Empria honshuana Prous & Heidemaa, sp. n., and Empria takeuchii Prous & Heidemaa, sp. n. The lectotypes of Poecilosoma pallipes Matsumura, 1912, Empria itelmena Malaise, 1931, Tenthredo candidata Fallén, 1808, and Tenthredo (Poecilostoma) hybrida Erichson, 1851 are designated. Empria itelmena Malaise, 1931, syn. n. is synonymized with Empria plana (Jakowlew, 1891). Poecilosoma pallipes Matsumura, 1912, previously assigned to Empria, is transferred to Monsoma, creating Monsoma pallipes (Matsumura, 1912), comb. n. Results of phylogenetic analyses using mitochondrial (COI) and nuclear (ITS1 and ITS2) sequences are also provided.