核糖体DNA ITS区序列在植物分子系统学研究中的价值

本文就近年来国内外有关被子植物核糖体ＤＮＡ中的内转录间隔区（ＩＴＳ）序列在植物属内,近缘属间乃至科内系统发育研究中的应用,结合作者在中国姜科山姜属（Ａｌｐｉｎｉａ Ｒｏｘｂ．）上的研究,对ＩＴＳ区序列在植物分子系统学研究中的价值作一简要综述,对其应用前景也进行了讨论。     

DNA序列在植物系统学研究中的应用

植物DNA序列由于进化速率上的差异而适用于不同分类阶元的系统发育研究,因此,针对某一特定的系统学问题选择相应合适的分子片段是分子系统学研究中最为关键的一步。在前人研究的基础上,主要讨论了目前分子系统发育和进化研究中一些常用的DNA序列的适用范围,包括nrDNA的18S基因及ITS等非编码区,cpDNA的编码基因（rbcI、matK、ndhF、atpB）及非编码区序列（rpL16、rpoC1、rps16、trnL-F和trnT-L）和应用较少的mtDNA。研究表明,18S、rbcI等编码基因及mtDNA一般适用于较高分类阶元甚至整个种子植物谱系间的系统发育的探讨,而ITS及cpDNA的非编码区序列等因其较快的进化速率多用于较低分类阶元的系统关系研究。     

地黄属植物的DNA条形码研究

地黄属(Rehmannia)为玄参科(Scrophulariaceae)药用植物,广泛分布于中国中东部及北部地区。由于地黄属植物经历了快速成种,导致其属内物种间形态性状差异较小,运用传统的形态学分类方法已难以准确地鉴定物种,近年来迅速发展起来的DNA条形码技术为快速、准确地鉴别物种提供了新思路。本研究选用3个叶绿体DNA非编码区片段(trn L-trn F、trn M-trn V和trn S-trn G)及核基因ITS片段,运用PWG-distance和TreeBuilding两种方法对地黄属5个物种75个个体进行了DNA条形码分析。结果表明:单个叶绿体DNA片段或核基因ITS片段对地黄属物种的鉴别率较低(0%~20%),组合的叶绿体DNA片段分辨能力虽然高于单个DNA片段,但并不能将地黄属5个物种完全区分开;trn S-trn G+ITS片段组合的分辨率可达100%,能够将地黄属5个物种准确区分,与所有叶绿体DNA片段和核基因ITS片段组合(trn L-trn F+trn M-trn V+trn S-trn G+ITS)的辨别率相同,因此推荐trn S-trn G+ITS作为地黄属植物的标准条形码。此外,利用DNA条形码鉴别物种时,可采用叶绿体DNA片段和核DNA片段组合的方法来提高物种鉴定的成功率。     

四个DNA 片段在山茶属分子系统学研究中的应用

选取山茶属14 个组中的10 组21 种植物对目前常用于属内种间的4 个DNA 片段( ITS、waxy 、trnLF
、rpL16) 进行序列测定。结果表明: ( 1) 来自叶绿体基因组的2 个片段( trnL- F、rpL 16) 其PCR 扩增和
测序都很容易, 但两者的进化速率都非常慢, 序列矩阵只有很少信息位点( trnL-F 含9 个, rpL16 为20
个) , 不能提供必要的系统发育信息。( 2) 来自核基因组的ITS 片段其PCR 产物比较容易获得, 但其序列
的测定存在较多问题。( 3) waxy 是来自核基因组的另一个片段, 其PCR 扩增因受模板DNA 的数量和质量
的影响很大而有一定难度, 但其进化速率较快, 序列矩阵具有较多信息位点( 92 个) , 并且在山茶属是单
拷贝, 这对于解决山茶属这类具有许多近缘物种的类群的系统关系有重要价值。基于tsnL- F、rpL16 和
waxy 三组数据所建分子系统树支持山茶属为一单系, 但属下系统由于取样等原因有待进一步研究。     

DNA序列在苔藓分子系统学研究中的应用

DNA是遗传信息的载体,直接对DNA核苷酸排列顺序的分析和比较是研究苔藓分子系统学的最彻底和最理想的方法。本文综述了DNA序列(叶绿体基因组、核基因组和线粒体基因组)在苔藓分子系统学中的应用,探讨了基因片段的选择策略。并提出只有将分子数据和传统分类学取得的研究成果结合起来,构建最合理的系统树,才能更好地推动苔藓分子系统学的发展。     

DNA序列在蕨类分子系统学研究中的应用

在分子系统学研究中,目的基因或者基因片段的选择是最关键的一步,由于进化速率的差异,不同的DNA序列适用于不同分类阶元的系统发育研究.本文综述了目前蕨类分子系统发育研究中常用的DNA序列分析,它们分别来自叶绿体基因组、核基因组和线粒体基因组,着重阐明叶绿体基因在蕨类分子系统学研究中的应用.本文还简要介绍了分子系统学研究中常见的问题及解决方法(如内类群和外类群的选择.适宜DNA片段的选择策略),总结了目前蕨类植物分子系统学研究所取得的进展和研究现状,展望了当今国际蕨类分子系统学的研究趋势.     

DNA 序列在蕨类分子系统学研究中的应用

在分子系统学研究中, 目的基因或者基因片段的选择是最关键的一步, 由于进化速率的差异, 不同的DNA序列适用于不同分类阶元的系统发育研究。本文综述了目前蕨类分子系统发育研究中常用的DNA序列分析, 它们分别来自叶绿体基因组、核基因组和线粒体基因组, 着重阐明叶绿体基因在蕨类分子系统学研究中的应用。本文还简要介绍了分子系统学研究中常见的问题及解决方法(如内类群和外类群的选择, 适宜DNA片段的选择策略), 总结了目前蕨类植物分子系统学研究所取得的进展和研究现状, 展望了当今国际蕨类分子系统学的研究趋势。     

叶绿体DNA及其在植物系统学研究中的应用

叶绿体ＤＮＡ及其在植物系统学研究中的应用黄瑶,李朝銮,马诚,吴乃虎（中国科学院成都生物研究所,成都,６１００４１）（中国科学院发育生物学研究所,北京,１０００８０）ＣＨＬＯＨＯＰＬＡＳＴＤＮＡＡＮＤＩＴＳＡＰＰＬＩＣＡＴＩＯＮＴＯＰＬＡＮＴＳＹＳＴＥ...     

部分百里香属植物分子系统学研究

采用ITS序列分析和ISSR分子标记技术,对中国野生百里香(5个种2个变种)及国外百里香(2个种)共计20份材料进行了属内种间及种下水平的系统发育和亲缘关系研究.结果表明:(1)ITS序列的系统发育树能较好地说明百里香属种间亲缘关系,地椒、地椒亚洲变种、百里香和Thymus serpyllum共同聚为一组,Bootstrap支持率为96%,表现为较近的亲缘关系;(2)基于ITS序列的系统发育树和ISSR多态性标记的聚类分析均将地椒怀远居群和地椒展毛变种兴凯居群聚为单独的一组,表现出与其它供试材料间较为明显的遗传差异;(3)ISSR遗传多态性分析将地椒、地椒亚洲变种和百里香很好的聚类;种下居群间的聚类则体现了与地理分布之间一定的相关性.将本实验结果与野外考查及形态学比较相结合,可以得出以下结论:地椒、地椒亚洲变种和百里香三者之间属于近缘种,并与国外的T.serpyllum种间有很近的亲缘关系;支持《黑龙江植物志》将地椒展毛变种兴凯居群列为单独的一个种;地椒怀远居群在遗传背景及分布生境上有很大的特殊性,对其在百里香属植物中的系统地位仍需进一步研究确定.     

四个DNA片段在山茶属分子系统学研究中的应用

选取山茶属14个组中的10组21种植物对目前常用于属内种间的4个DNA片段（ITS、waxy、trnL-F、rpL16）进行序列测定。结果表明：（1）来自叶绿体基因组的2个片段（trnL-F、rpL16）其PCR扩增和测序都很容易,但两者的进化速率都非常慢,序列矩阵只有很少信息位点（trnL-F含9个,rpL16为20个）,不能提供必要的系统发育信息。（2）来自核基因组的ITS片段其PCR产物比较容易获得,但其序列的测定存在较多问题。（3）waxy是来自核基因组的另一个片段,其PCR扩增因受模板DNA的数量和质量的影响很大而有一定难度,但其进化速率较快,序列矩阵具有较多信息位点（92个）,并且在山茶属是单拷贝,这对于解决山茶属这类具有许多近缘物种的类群的系统关系有重要价值。基于tsnL-F、rpL16和waxy三组数据所建分子系统树支持山茶属为一单系,但属下系统由于取样等原因有待进一步研究。     

Are nuclear loci ideal for barcoding plants? A case study of genetic delimitation of two sister species using multiple loci and multiple intraspecific individuals

Considerable debate remains as to which DNA region should be used to barcode plants. Several different chloroplast (cp) DNA regions (rbcL, matK, and trnH-psbA) and nuclear ribosomal internal transcribed regions (ITS) have been suggested as suitable barcodes in plants. Recently, low-copy nuclear loci were also suggested to be potentially ideal barcode regions. The aim of the present study was to test the effectiveness of these proposed DNA fragments and five additional low-copy loci (CHS, DETl, COPl, PGICl, and RPS2; comprising both coding and non-coding regions) in barcoding closely related species. We examined the divergences within and between two species of Pugioniun (Brassicaceae). We failed to find any interspecific variation from three cpDNA fragments with which to discriminate the two species. However, a single base mutation in the internal transcribed spacer (ITS) could discriminate between the two species consistently. We found more variations among all individuals of the two species using each of the other five low-copy nuclear loci. However, only alleles from one locus (DET1) of the five low-copy loci related to flowering regulations was able to distinguish the sampled individuals into two species. We failed to amplify the corresponding fragments out of Brassicaceae using the designed DETl primers. We further discussed the discrimination power of different loci due to incomplete lineage sorting, gene flow, and species-specific evolution. Our results highlight the possibility of using the nuclear ITS as a core or complementary fragment to barcode recent diverged species.     

Are nuclear loci ideal for barcoding plants? A case study of genetic delimitation of two sister species using multiple loci and multiple intraspecific individuals

Considerable debate remains as to which DNA region should be used to barcode plants. Several different chloroplast (cp) DNA regions (rbcL, matK, and trnH–psbA) and nuclear ribosomal internal transcribed regions (ITS) have been suggested as suitable barcodes in plants. Recently, low-copy nuclear loci were also suggested to be potentially ideal barcode regions. The aim of the present study was to test the effectiveness of these proposed DNA fragments and five additional low-copy loci (CHS, DET1, COP1, PGIC1, and RPS2; comprising both coding and non-coding regions) in barcoding closely related species. We examined the divergences within and between two species of Pugionium (Brassicaceae). We failed to find any interspecific variation from three cpDNA fragments with which to discriminate the two species. However, a single base mutation in the internal transcribed spacer (ITS) could discriminate between the two species consistently. We found more variations among all individuals of the two species using each of the other five low-copy nuclear loci. However, only alleles from one locus (DET1) of the five low-copy loci related to flowering regulations was able to distinguish the sampled individuals into two species. We failed to amplify the corresponding fragments out of Brassicaceae using the designed DET1 primers. We further discussed the discrimination power of different loci due to incomplete lineage sorting, gene flow, and species-specific evolution. Our results highlight the possibility of using the nuclear ITS as a core or complementary fragment to barcode recent diverged species.     

Phylogeny of the New World diploid cottons (Gossypium L., Malvaceae) based on sequences of three low-copy nuclear genes

American diploid cottons (Gossypium L., subgenus Houzingenia Fryxell) form a monophyletic group of 13 species distributed mainly in western Mexico, extending into Arizona, Baja California, and with one disjunct species each in the Galapagos Islands and Peru. Prior phylogenetic analyses based on an alcohol dehydrogenase gene (AdhA) and nuclear ribosomal DNA indicated the need for additional data from other molecular markers to resolve phylogenetic relationships within this subgenus. Toward this end, we sequenced three nuclear genes, the anonymous locus A1341, an alcohol dehydrogenase gene (AdhC), and a cellulose synthase gene (CesA1b). Independent and combined analyses resolved clades that are congruent with current taxonomy and previous phylogenies. Our analyses diagnose at least two long distance dispersal events from the Mexican mainland to Baja California, following a rapid radiation of the primary lineages early in the diversification of the subgenus. Molecular data support the proposed recognition of a new species closely related to Gossypium laxum that was recently collected in Mexico.     

利用低拷贝核基因重建菊科紫菀亚科族间系统发育关系

紫菀亚科(Asteroideae)是菊科最大的一个亚科, 包含的种数多于被子植物的绝大多数科。目前, 紫菀亚科族间的系统发育关系主要依赖于叶绿体基因信息, 但是叶绿体基因为单亲遗传, 并不能完整反映进化历史。鉴于杂交现象在菊科普遍存在, 故利用核基因可以反映更完整的紫菀亚科进化历史。该研究首次使用从转录组数据(20个新测+11个从NCBI数据库下载)中筛选出的47个直系同源低拷贝核基因来研究紫菀亚科的系统发育关系, 共选取了29个物种, 代表了紫菀亚科20个族中的13个族。用超矩阵分析方法和溯祖推测分析方法各获得了1个稳定的紫菀亚科系统树, 每个树上绝大多数分支都得到了高度支持, 且2个树之间没有明显的冲突。新的紫菀亚科族间系统发育关系揭示了千里光超族应并入紫菀超族, 春黄菊族可能是千里光族与紫菀族杂交起源的, 金鸡菊族很可能也是杂交起源的。该研究结果显示低拷贝核基因可以更好地解决科以下分类阶元的系统发育关系, 对菊科乃至被子植物其它科的系统发育研究具有重要的借鉴意义。     

Phylogeny and biogeography of the flowering plant genus Styrax (Styracaceae) based on chloroplast DNA restriction sites and DNA sequences of the internal transcribed spacer region

Phylogenetic relationships within the flowering plant genus Styrax were investigated with DNA sequence data from the internal transcribed spacer (ITS) region of nuclear ribosomal DNA (nrDNA) and with chloroplast DNA restriction site data from the genes trnK, rpoC1, and rpoC2. The data sets from each genome were analyzed separately and in combination with parsimony methods. The results strongly support the monophyly of each of the four series of the genus but provide little phylogenetic resolution among them. Reticulate evolution may at least partly explain discordance between the molecular phylogenetic estimates and a prior morphological estimate within series Cyrta. The historical biogeography of the genus was inferred with unweighted parsimony character optimization of trees recovered from a combined ITS and morphological data set, after a series of combinability tests for data set congruence was conducted. The results are consistent with the fossil record in supporting a Eurasian origin of Styrax. The nested phylogenetic position of the South American members of the genus within those from southern North America and Eurasia suggests that the boreotropics hypothesis best explains the amphi-Pacific tropical disjunct distribution occurring within section Valvatae. The pattern of relationship recovered among the species of section Styrax ((western North America + western Eurasia) (eastern North America + eastern Eurasia)) is rare among north-temperate Tertiary forest relicts. The monophyly of the group of species from western North America and western Eurasia provides qualified support for the Madrean-Tethyan hypothesis, which posits a Tertiary floristic connection among the semiarid regions in which these taxa occur. A single vicariance event between eastern Asia and eastern North America accounts for the pattern of relationship among intercontinental disjuncts in series Cyrta.     

黑龙江悬钩子属植物叶表皮形态结构的研究

利用扫描电子显微镜对黑龙江悬钩子属植物的叶表皮形态结构进行比较研究。结果显示:(1)悬钩子属植物叶的上表皮细胞呈多边形,垂周壁平直,或无规则形,垂周壁浅波纹;下表皮细胞无规则形,垂周壁浅波纹或深波纹。(2)表皮毛类型有单细胞直立不分支、卷曲不分支,头状腺毛和盾状腺毛四种类型。(3)气孔器均分布于下表皮,且气孔器类型为无规则形;气孔外拱盖单层、内缘平滑或不规则波状。研究表明,黑龙江悬钩子属植物的叶表皮微形态学特征表现出一定差异性,对种间的划分和鉴定具有一定的分类学意义。     

Systematic overview of Krigia (Asteraceae-Lactuceae)

Cytological and morphological variation among Krigia species is examined. Krigia exhibits a broad range of chromosome numbers including n = 4, 5, 6, 9, 10, 15, and 30. Section Krigia is characterized by reflexed phyllaries and a base chromosome number of x = 5, while section Cymbia is characterized by erect phyllaries and chromosome numbers of n = 4, 6, and 9. The micromorphological characteristics of achenes, pappus, styles, corolla, pollen, stomata, and trichomes are documented using scanning electron microscopy. Among these, the pappus shows the greatest diversity and three major types are identified: 1) a pappus of many bristles and scales, as in K. dandelion, K. montana, and K. biflora; 2) a pappus of five bristles and five scales, as in K. virginica and K. occidentalis; and 3) an absent or highly reduced pappus, as in the K. cespitosa complex and K. wrightii. Thirty-five cytological and morphological characters are subjected to phylogenetic analyses. The two sections, Krigia and Cymbia, form monophyletic lineages. Within section Krigia, the annual species, K. virginica, forms an independent clade, while the perennial species, K. dandelion, K. biflora, and K. montana, form a monophyletic clade. Krigia montana and K. biflora are identified as sister species and a hybrid between these has been identified. The hybrid is more similar morphologically to K. montana than K. biflora. Within section Cymbia, phylogenetic relationships among K. wrightii, K. occidentalis, and K. cespitosa are uncertain. Nine taxa of Krigia are herein recognized: K. dandelion, K. biflora, K. biflora var. viridis (comb. nov.), K. montana, K. virginica, K. wrightii (comb. nov.), K. occidentalis, K. cespitosa, and K. cespitosa f. gracilis (comb. nov.). Phylogenetic relationships among 12 taxa of Krigia species are compared using various combinations of morphology, chloroplast DNA, and nuclear ribosomal DNA data. Tree topologies from different combinations of data are largely congruent. The most resolved phylogenetic tree is obtained using the combined data from morphology, chloroplast DNA, and nuclear ribosomal DNA.     

Chloroplast genes encoding subunits of the H+-ATPase complex of Chlamydomonas reinhardtii are rearranged compared to higher plants: sequence of the atpE gene and location of the atpF and atpI genes

Summary The chloroplast gene for the epsilon subunit (atpE) of the CF₁/CF₀ ATPase in the green alga Chlamydomonas reinhardtii has been localized and sequenced. In contrast to higher plants, the atpE gene does not lie at the 3 end of the beta subunit (atpB) gene in the chloroplast genome of C. reinhardtii, but is located at a position 92 kb away in the other single copy region. The uninterrupted open reading frame for the atpE gene is 423 bp, and the epsilon subunit exhibits 43% derived amino acid homology to that from spinach. Codon usage for the atpE gene follows the restricted pattern seen in other C. reinhardtii chloroplast genes.The genes for the CF₀ subunits I (atpF) and IV (atpI) of the ATPase complex have also been mapped on the chloroplast genome of C. reinhardtii. The six chloroplast ATPase genes in C. reinhardtii are dispersed individually between the two single copy regions of the chloroplast genome, an organization strikingly different from the highly conserved arrangement in two operon-like units seen in chloroplast genomes of higher plants.Abbreviations bp base pairs - CF₁ chloroplast coupling factor 1 - CF₀ chloroplast coupling factor 0 - F₁ coupling factor 1 - F₀ coupling factor 0 - kb kilobase pairs