草兔线粒体基因组全序列及其系统发育关系分析

草兔(Lepus capensis)在我国数量多,分布广,但人们对草兔的系统地理学、草兔与其他兔类的系统发育关系了解不多,其亚种水平的分类也长期存在争议。本研究通过LA-PCR 技术对草兔线粒体基因组全序列进行PCR 扩增、序列测定和分析,并基于线粒体基因组12 个蛋白质编码基因10 776 bp的核苷酸序列,采用贝叶斯法(BI)和最大似然法(ML)构建哺乳纲(Mammalia) 灵长总目(Euarchontoglires)5 个目12 种动物的系统发育关系,结果支持兔形目(Lagomorpha)的单系起源,其中草兔与欧洲野兔(L. europaeus)亲缘关系最近,两者互为姐妹群,进而与家兔(Oryctolagus cuniculus)构成姐妹群关系。此外,基于线粒体细胞色素b 基因全序列,构建31 个兔类个体的系统发育关系,结果显示,草兔与雪兔(L. timidus),云南兔(L. comus) 与高原兔(L.oiostolus),东北兔(L. mandschuricus)与塔里木兔(L. yarkandensis) 有较近的亲缘关系,而在草兔内部,中国的草兔与南非草兔亲缘关系较远,采自江苏盐城的草兔与来自山东、陕西、四川的草兔聚为一支,具有明显较近的亲缘关系。该研究为更好地探讨草兔系统发育关系提供了进一步的分子生物学资料.     

青蟹线粒体COI假基因的分离和特征分析

线粒体DNA标记在遗传结构和系统进化研究中得到广泛应用,然而核假基因的存在对此有很大威胁。本文以中国东南沿海的青蟹(Scylla paramamosain)为研究对象,利用线粒体COI基因的通用引物和特异性引物进行扩增,分别得到34个假基因（nuclear mitochondrial pseudogenes, Numts）和5个线粒体COI基因序列。在所获得的34个假基因中共定义了29种单倍型,根据序列的相似度,这些假基因可以分为2类,每类假基因都有各自保守的核苷酸序列。第Ⅰ类假基因存在2处插入序列和1处8 bp的缺失序列,这些位点导致了整个阅读框的移位;在第Ⅱ类假基因和5个线粒体COI序列中只有碱基替换,未发现插入和缺失序列。实验结果分析表明,这两类假基因分别代表了2次核整合事件,即核转移事件的最低值。研究结果提示了     

基于线粒体COI基因全序列的直翅目部分类群系统发育关系分析

该研究基于直翅目56种昆虫的COI基因全序列构建了该目部分类群间的系统发育关系,同时也分析了COI基因编码的氨基酸序列构建直翅目系统发育关系的可靠性。将COI序列按照密码子一、二、三位点划分,分别计算PBS(partioned Bremer support)值,评估蛋白质编码基因密码子不同位点的系统发生信号强度。分析结果支持螽亚目和蝗亚目的单系性;剑角蝗科、斑腿蝗科、斑翅蝗科、网翅蝗科和槌角蝗科5科均不是单系群,科间的遗传距离在0.107~0.153之间变化,与其他科相比遗传距离较小,符合将这5科合并为一科(即蝗科)的分类系统,瘤锥蝗科和锥头蝗科归为锥头蝗总科,癞蝗科单独成为一科,这也与Otte(1997)系统的划分一致。根据PBS值的大小推断密码子第三、第一位点对系统树分支的贡献比第二位点大,并且较长的序列含有较多的信息位点。研究也证实将各物种COI基因之间的遗传距离作为直翅目划分科级阶元的工具是可行的。     

鳜类系统发育的线粒体Cytb基因全序列分析

测定了鳜、大眼鳜、斑鳜、暗鳜、波纹鳜、长体鳜、中国少鳞鳜等7种鳜类12个个体的线粒体细胞色素b基因全序列。结合GenBank中的同源序列,共分析了9种鳜类的系统发育关系。序列分析表明,鳜属鱼类属内种间的遗传距离(0.015～0.093)明显小于少鳞鳜属鱼类属内种间的遗传距离(0.152～0.178)。在分子系统发育树上,长体鳜与鳜属的鳜、大眼鳜、斑鳜、波纹鳜、暗鳜聚合成一分支,少鳞鳜属的种类聚成另一分支;支持将长体鳜归入鳜属,鳜类分为鳜属和少鳞鳜属等二个属的分类处理。在鳜属鱼类中,鳜和大眼鳜亲缘关系十分密切;斑鳜与波纹鳜亲缘较近;长体鳜与鳜属其它5个种的亲缘关系较远。在少鳞鳜属鱼类中,中国少鳞鳜和日本少鳞鳜的亲缘关系较远,韩国少鳞鳜的系统位置较不明确。鳜类的单系性及其鳜类的系统位置仍有待进一步研究。     

基于线粒体细胞色素c氧化酶亚基I基因序列的帘蛤科贝类分子系统发育研究

对21种帘蛤科贝类线粒体细胞色素c氧化酶亚基Ⅰ(cytochrome c oxidase subunit I,COI)基因核苷酸序列进行了分析,以探讨这一序列在种质鉴定、分子系统发生研究中的应用价值。测序结果表明,所有物种扩增片段长度均为707 bp(含引物),序列A+T含量(62.4%—67.8%)明显高于G+C含量。物种间共有变异位点379个,其中简约信息位点334个;此区段共编码235个氨基酸,种间共有氨基酸变异位点100个。以COI基因片段序列为标记,用中国蛤蜊(Mactra chinensis)作外群,构建了35种帘蛤科贝类(其中14种贝类COI序列从GenBank下载)的系统发生树,结合拓扑结构分析和序列比对分析,结果表明:支持将短文蛤(Meretrix petechinalis)和丽文蛤(M.lusoria)订为文蛤(M.meretrix)的同物异名的观点,建议将丽文蛤和短文蛤订为文蛤的地理亚种;支持将薄片镜蛤(Dosinia corrugata)和D.angulosa订为2个独立种的观点;认为将波纹巴非蛤(Paphia undulata)和织锦巴非蛤(P.textile)订为2个独立种是合适的。COI基因序列含有丰富的遗传信息,适合作为帘蛤科贝类种群遗传结构和系统发生研究的分子标记。     

家鹅品种细胞色素b基因序列分析及其系统发育关系

测定了15个中国家鹅和2个欧洲家鹅品种44个个体线粒体细胞色素b基因全序列（1143 bp）,并结合GenBank中的野生种白额雁序列比较分析表明,检测到31个可变位点和4种单倍型,没有发现缺失或插入,核苷酸多样度和单倍型多样度分别为0.00648和0.45,密码子第三位点G碱基的含量较低（14.2%）,A、T和C的含量极相似,第一、二位点的偏倚度（0.057和0.223）低于第三位点（0.492）,转换数明显高于颠换数(Ts/Tv＝9.5～19),密码子第三位点的转换数最高。系统发育分析结果支持中国家鹅的两个不同起源学说。     

基于线粒体细胞色素b基因序列的雀形目18种鸟系统发育关系

采用线粒体细胞色素b(cytochrome b)基因,利用DNA测序的方法研究了雀形目18种鸟类的分子系统发育关系。通过序列分析比对,运用邻接法和最大似然法构建了雀形目18种鸟类的系统发育树,结果表明,燕雀类和类的分歧程度达到了科级水平,建议分别独立成科;长尾山雀可从山雀科中分离出来,单列成科;柳莺归入柳莺亚科(Phylloscopinae),树莺归入大苇莺亚科(Acrocephalinae)。Cytb基因的各主要分支中存在着相对恒定的分子钟,根据鸟类mtDNA的cytb进化速度大约是每百万年1.6%,得到18种鸟的科间分歧时间在10.5百万年左右,科内分歧在9.0百万年左右。     

家鸭细胞色素C氧化酶Ⅲ（COⅢ）基因的克隆及序列分析

线粒体是存在于绝大多数真核细胞内的一种基本的、重要的细胞器,是细胞进行氧化磷酸化的场所。不同脊椎动物同源基因序列的比较显示,细胞色素b(Cytb)、细胞色素C氧化酶Ⅰ（COⅠ）、细胞色素C氧化酶Ⅱ（COⅡ）、细胞色素C氧化酶Ⅲ（COⅢ） 基因最保守,同源性最高,ATPase6、ATPase8基因、ND基因变异比较大。本试验以家鸭（家鸭起源于绿头鸭：Anas platyrhynchos）肝脏的线粒体DNA为模板,按照GenBank已经公布的潜鸭族（AF090337）的全序列及其绿头鸭的mtDNA部分序列（L22476、L16770、L22477）设计合成特异引物进行PCR扩增,克隆并测定了线粒体细胞色素C氧化酶Ⅲ亚基（COⅢ）的全序列784bp以及ATPase6基因的3'端和tRNA-Gly基因的5'端序列共934bp。 用DNAStar分析软件对家鸭与GenBank中7种禽类的COⅢ序列进行比较分析,显示家鸭与这些动物的COⅢ基因具有较高的同源性,与同科潜鸭属中的Aythya Americana的相似性 最高为90.6 %,与同目不同科的加拿大雁、小天鹅、白额雁的相似性分别为89%、88.6%、88.6%。根据家鸭与其他7种禽类的COⅢ基因序列相似性所建立的进化树,与传统的分类地位基本吻合。     

基于细胞色素b基因序列的东亚鳜类系统发育关系

鳜类为东亚特有类群。为验证东亚鳜类的单系起源,解决其分类划分的争议,本文采用特异性引物PCR扩增法获得了鳜类中鳜、大眼鳜、斑鳜、暗鳜、波纹鳜、长体鳜细胞色素b基因的全长序列,结合GenBank中日本少鳞鳜、朝鲜少鳞鳜和中国少鳞鳜的序列,初步构建了东亚鳜类种间的系统发育关系。NJ树和MP树均一致表明,鳜类为单系类群,可分为两支:鳜鱼群和少鳞鳜群。长体鳜未单独成群,而与鳜鱼属聚为一支,不支持将长体鳜单独设属,而应归入鳜鱼属。鳜鱼群中,MP树推测的长体鳜、暗鳜较为原始,鳜、大眼鳜、斑鳜和波纹鳜为特化种类的系统关系假设与其形态、生态特征表型进化较为一致。少鳞鳜群中,中国少鳞鳜与朝鲜少鳞鳜为姐妹种,再与日本少鳞鳜构成姐妹群的系统关系与其地理分布格局较为一致。本研究结果可为深入探讨鳜类物种有效性、系统分类位置及生物地理学提供依据。     

基于线粒体基因Cytb和COI的蝗科中五亚科分子系统发育关系分析（直翅目: 蝗总科）（英文）

本研究基于Cytb 基因和COI基因的部分序列来推断17种蝗虫之间的系统发育关系。这17种蝗虫均采自国内,代表了蝗科(Acrididae)5个亚科：黑蝗亚科(Melanoplinae)、斑腿蝗亚科(Catantopinae)、刺胸蝗亚科(Cyrtacanthacridinae)、斑翅蝗亚科(Oedipodinae)和大足蝗亚科(Gomphocerinae)。采用联合序列方法进行分析,结果显示：Cytb 和COI联合序列长度为1 998 bp,其中A和T总含量为72.13%,G和C总含量为27.87%。联合序列共包含了889个保守位点,1 109个变异位点,在这些变异位点中有838个简约信息位点。系统发生树采用邻接法（NJ）、最大简约法（MP）和最大似然法（ML）进行构建。使用蜢总科的变色乌蜢Erianthus versicolor 和 Erianthus sp. 两个种作为外群。结果表明：大足蝗亚科和斑腿蝗亚科的单系性没有得到支持。斑翅蝗亚科内部各种聚成一个大支,在本研究中该亚科的单系性得到支持,与前人的研究结论相同。大足蝗亚科、斑腿蝗亚科、刺胸蝗亚科和黑蝗亚科这4科关系非常近,可以考虑将其合并为一个亚科。同时,我们发现基于Cytb和COI基因联合序列推断蝗科内各亚科间的系统发生关系并不十分可靠。     

The origin of the Tibetan Mastiff and species identification of Canis based on mitochondrial cytochrome c oxidase subunit I (COI) gene and COI barcoding

DNA barcoding is an effective technique to identify species and analyze phylogenesis and evolution. However, research on and application of DNA barcoding in Canis have not been carried out. In this study, we analyzed two species of Canis, Canis lupus (n = 115) and Canis latrans (n = 4), using the cytochrome c oxidase subunit I (COI) gene (1545 bp) and COI barcoding (648 bp DNA sequence of the COI gene). The results showed that the COI gene, as the moderate variant sequence, applied to the analysis of the phylogenesis of Canis members, and COI barcoding applied to species identification of Canis members. Phylogenetic trees and networks showed that domestic dogs had four maternal origins (A to D) and that the Tibetan Mastiff originated from Clade A; this result supports the theory of an East Asian origin of domestic dogs. Clustering analysis and networking revealed the presence of a closer relative between the Tibetan Mastiff and the Old English sheepdog, Newfoundland, Rottweiler and Saint Bernard, which confirms that many well-known large breed dogs in the world, such as the Old English sheepdog, may have the same blood lineage as that of the Tibetan Mastiff.     

Identification of forensically important blow fly species (Diptera: Calliphoridae) in China by mitochondrial cytochrome oxidase I gene differentiation

Abstract Unambiguous and rapid sarcosaphagous insect species identification is an essential requirement for forensic investigations. Although some insect species are difficult to classify morphologically, they can be effectively identified using molecular methods based on similarity with abundant authenticated reference DNA sequences in local databases. However, local databases are still relatively incomplete in China because of the large land area with distinct regional conditions. In this study, 75 forensically important blow flies were collected from 23 locations in 16 Chinese provinces, and a 278‐bp segment of the cytochrome oxidase subunit I gene of all specimens was successfully sequenced. Phylogenetic analysis of the sequenced segments showed that all Calliphorid specimens were properly assigned into nine species with relatively strong supporting values, thus indicating that the 278‐bp cytochrome oxidase subunit one region is suitable for identification of Calliphorid species. The clear difference between intraspecific threshold and interspecific divergence confirmed the potential of this region for Calliphorid species identification, especially for distinguishing between morphologically similar species. Intraspecific geographic variations were observed in Lucilia sericata (Meigen, 1826) and Lucilia caesar (Linnaeus, 1758).     

Population genetic diversity of Chinese sea bass (Lateolabrax maculatus) from southeast coastal regions of China based on mitochondrial COI gene sequences

Chinese sea bass (Lateolabrax maculatus), an important commercial and recreational fishery species in China, whose natural resources have decreased dramatically for decades due to overfishing and environmental changes. To provide guidelines for the source conservation and management, genetic diversity and population structure of L. maculatus were analyzed based on the mitochondrial cytochrome oxidase subunit I (COI) gene. Totally 192 individuals were sampled from five locations: Qingdao (QD), Chongming (CM), Dongtou (DT), Lieyu (LY) and Fangcheng (FC). After sequencing of a 586 bp fragment of COI gene, 20 haplotypes were defined. H4 (haplotype 4) and H8 were the dominant haplotypes and existed in all populations. Haplotype diversity (Hd) and nucleotide diversity (π) of each population ranged from 0.572 to 0.721 and from 0.00129 to 0.00271, respectively. The highest Hd and π were found in DT and LY populations, while the lowest value of Hd and π were in the QD population. Genetic distance ranged from 0.0031 to 0.0027 within populations and from 0.0018 to 0.0035 between populations. Analysis of molecular variance (AMOVA) revealed that significant genetic divergence was found in QD and FC populations. In addition, neutrality tests and mismatch distribution analysis indicated that this species experienced potential population expansion events.     

Phylogenetic analysis of six species of pacific abalone (Haliotidae) based on DNA sequences of 16s rRNA and cytochrome c oxidase subunit I mitochondrial genes

Six species of abalones (Haliotidae) are found on the Korean coasts. Identification and characterization of these abalones are usually based on morphologic characters. In this research we compared the partial sequences of the mitochondrial 16S ribosomal RNA and cytochrome c oxidase subunit I genes to identify species using molecular data and to determine their phylogenetic relationships. Sequence alignments and phylogenetic analysis revealed that the 6 species fell into 2 distinct groups which were genetically distant from each other and exhibited little internal phylogenetic resolution. One group included Haliotis discus hannai, H. discus discus, H. madaka, and H. gigantea, while the other group contained H. diversicolor supertexta and H. diversicolor diversicolor. The 16S rRNA sequences were relatively more conserved than to the COI sequences, but both gene sequences provided sufficient phylogenetic information to distinguish among the 6 species of Pacific abalone, and thus could be valuable molecular characters for species identification.     

基于Cytb和COⅠ基因部分序列的15种麻蝇之间的系统发育关系(双翅目：麻蝇科)

本研究通过测序Cytb基因和COⅠ基因的部分序列来推定15种麻蝇之间的系统发育关系。在世界麻蝇名录中,本研究的15种麻蝇能够代表麻蝇属Sarcophaga的6个亚属。连接序列（972 bp）被用于系统发育分析;分析方法包括了了最大简约法、最大似然法以及贝叶斯法。我们的结果提示了亚麻蝇亚属Parasarcophaga、别麻蝇亚属Boettcherisca以及红麻蝇亚属Liopygia的单系性,同时也表明蛇麻蝇亚属Liosarcophaga和德麻蝇亚属Pandelleisca并不是单源的。不过,目前的研究并不能分辨野德麻蝇S. (Pandelleisca) similis和峨眉叉麻蝇S. (Robineauella) coei的系统发育位置。此外,最大简约分析和似然功能分析在scopariiformis-iwuensis进化枝和polystylata-hui进化枝的关系上产生了不一致的系统发育推断。因此,后续研究不仅需要其他的分子标记,也需要更多的分类取样。     

Effects of habitat fragmentation on genetic diversity and gene flow among the Homidia socia (Collembola: Entomobryidae) populations in the Thousand Island Lake

In the present study, partial sequences of the mitochondrial cytochrome oxidase subunit I (COI) gene of 22 island populations of the springtail Homidia socia in the Thousand Island Lake were sequenced. Across all sequences, 37 haplotypes were identified for the 510‐bp mitochondrial (mt) DNA COI gene. Haplotype 2 was the most common, and was distributed in the most of the 22 island populations. Haplotype diversity ranged from 0.065 to 0.733, and the total genetic diversity was 0.56216. The genetic characteristics of the 22 island populations were analyzed using the fixation index and gene flow, with values of 0.00043–0.94900 and 0.02703–703.72540, respectively. Comparison between (island area and isolations) with population genetic diversity revealed that there were no significant correlations between them, except for a significant correlation between the number of haplotypes and island area. Mantel tests showed that there was no significant correlation between geographic distance and genetic distance among various groups. All the results indicated that there were no obvious relationships between island characteristics and the genetic diversity of the springtails. We consider that the low dispersal capacity of springtails and the island patches surrounded by water in the Thousand Island Lake are the major factors affecting the genetic diversity of H. socia.     

Identification of mosquito bloodmeals using mitochondrial cytochrome oxidase subunit I and cytochrome b gene sequences

Abstract.  Primer pairs were designed and protocols developed to selectively amplify segments of vertebrate mitochondrial cytochrome oxidase subunit 1 (COI) and cytochrome b (Cyt b ) mtDNA from the bloodmeals of mosquitoes (Diptera: Culicidae). The protocols use two pairs of nested COI primers and one pair of Cyt b primers to amplify short segments of DNA. Resultant sequences are then compared with sequences in GenBank, using the BLAST function, for putative host identification. Vertebrate DNA was amplified from 88% of our sample of 162 wild-caught, blood-fed mosquitoes from Oregon, U.S.A. and GenBank BLAST searches putatively identified 98% of the amplified sequences, including one amphibian, seven mammalian and 14 avian species. Criteria and caveats for putative identification of bloodmeals are discussed.     

基于线粒体 COI 和 Cytb 基因的中国灰蝶三亚科主要类群间的系统发育关系分析

【目的】针对中国灰蝶科中亲缘关系较近的3个主要亚科［灰蝶亚科(Lycaeninae)、线灰蝶亚科(Theclinae)以及眼灰蝶亚科(Polyommatina)］,基于线粒体基因序列数据研究它们主要类群间的系统发育关系。【方法】对3亚科共53种灰蝶的线粒体 COI 和 Cytb 基因进行序列测定和序列变异分析,同时,基于最大似然法(maximum likelihood, ML)和贝叶斯法(bayesian inference, BI)等建树方法重建53种灰蝶的系统发育树。【结果】串联的2个基因共1 431 bp,其中保守位点855个,可变位点576个,简约信息位点488个;A+T的平均含量为74.5%,明显高于G+C的平均含量(25.5%)。系统树显示,灰蝶亚科以及眼灰蝶亚科均是单系发生,线灰蝶亚科则为并系群。全部灰蝶物种共分为三大支系：灰蝶亚科为第1支系;眼灰蝶亚科与线灰蝶亚科中的旖灰蝶族(Hypolycaenini)、富丽灰蝶族(Aphnaeini)分别构成单系群并互为姊妹群,它们共同构成第2支系;线灰蝶亚科中的美灰蝶族(Eumaeini)、玳灰蝶族(Deudorigini)、娆灰蝶族(Arhopalini)和线灰蝶族(Theclini)构成第3支系,其亲缘关系为：（（（线灰蝶族+娆灰蝶族）+玳灰蝶族）+美灰蝶族）。【结论】本研究涉及的3个灰蝶亚科中,灰蝶亚科是一个独立的支系,眼灰蝶亚科与线灰蝶亚科之间有较近的亲缘关系,但它们内部主要类群间的系统发育关系还需要进一步的研究。     

Identification of elmid larvae (Coleoptera: Elmidae) from Sanin District of Honshu,Japan, based on mitochondrial DNA sequences

Larvae of Elmidae from the Sanin District, Honshu, Japan, were classified into 14 types based on morphological features, of which 11 types were unidentified for species. Species or genus of the unidentified types were determined by comparing their mitochondrial cytochrome oxidase subunit I gene sequences with those of identified adult specimens. A new key to species/genera of elmid larvae was proposed.