Nucleotide composition and contents of CpG and CpNpG in the ITS1, ITS2 and 5.8S rRNA of the representatives of phylogenetic branches of melanthiales-liliales and melanthiales and melanthiales-asparagales (angiospermae, monocotyledones) reflects a characteristic feature of evolution

Variations of nucleotide composition and frequency of CpG and CpNpG sequences in the clusters of nuclear ribosomal genes of taxa, belonging to two phylogenetic branches of Angiospermae have been statistically analyzed. This region of eucaryotic genomes is nucleolus organizer and functions in a separate compartment of cell nucleus that can do running here processes it is enough specific. It is shown that level of evolution advance of a taxon, defined on morphological data, is in positive correlation with quantitative value of dC, CpG and CpNpG. This is found in contradiction with beliefs about the general rules of the transformation of nucleotide composition in evolution, that suggest a CpG suppression. Cryptaffinous taxa being connecting link between a well morphological outlined taxonomic group, differ to be raised in     

Application of ITS sequences of nuclear rDNA in phylogenetic and evolutionary studies of angiosperms

Nuclear rRNA genes (rDNA) in angiosperms are arranged in long tandem repeat ing units, much like those of other higher eukaryotes. Owing to rapid concerted evolution, the repeat units have homogenized or nearly so in most species. The internal transcribed spacer (ITS) of nuclear rDNA is composed of ITS1 and ITS2, which are seperated by 5.8S rDNA. The two spacers, ITS1 (187～298 bp) and ITS2 (187～252 bp), can be readily amplified by PCR and sequenced using universal primers. The sequences contain many vari able sites and potential informative sites among related species, and have been proven to be a useful molecular marker in phylogenetic and evolutionary studies of many angiosperm taxa. It can be used not only in classification and phylogenetic inferences at the levels of family, subfamily, tribe, genus and section, but also in reconstruction of reticulate evolution and de tection of the speciation via hybridization and polyploidization. But this region may not be useful for resolving phylogenetic relationships among families or taxa of higher hierarchy ow- ing to the rapid variation of the ITS sequences.     

基于ITS序列探讨山茶属金花茶组的系统发育关系

测定了分布于我国的 2 2个山茶属金花茶组的种或变种的nrDNAITS区序列 ,它们的序列长度在 476～ 496之间。GC含量都超过了 70 % ,应用Kimura2 模型计算了序列间的分化程度 ,构建了最大简约树、邻接树和最大似然树 ,分析结果表明 :( 1 )淡黄金花茶、毛籽金花茶、陇瑞金花茶、弄岗金花茶、大样金花茶和凹脉金花茶的关系较近 ;( 2 )小瓣金花茶、小花金花茶、薄叶金花茶、多瓣金花茶、夏石金花茶和龙州金花茶的关系较近。ITS区序列分析结果与AFLP分析结果相近     

A new insight into the phylogeny of vascular cryptogams with special reference to Selaginella and Isoetes inferred from nuclear ITS/5.8S rDNA sequences

In order to provide a better understanding of the evolutionary history of vascular cryptogams, phylogenetic framework was developed based on ITS1, ITS2 and 5.8S rDNA sequences of 102 extant taxa of vascular cryptogams using Maximum Parsimony (MP) analysis. The analysis revealed high GC content in Isoetaceae (60.5 %) in comparison with Selaginellaceae (54.4 %) that was envisaged to be the result of variation in selection, mutational bias, and biased recombination-associated DNA repair within these two plant lineages during evolution. Transition/transversion ratio was observed to be 0.9 in Isoetaceae, 0.68 in Selaginellaceae and 0.57 among all the 102 taxa belonging to lycophytes and ferns. It is hypothesized that the lycophytes have been separated very early during evolution and therefore acquired independent line of evolution from the other plant lineages. Although Selaginellaceae and Isoetaceae are closely related ancient plant groups, pairwise sequence divergence of sampled taxa on the basis of transition and transversion, and disparity index values per site between sampled sequence pairs pointed towards the differential investment of natural selection process. These lead to high rate of nucleotide substitution within nuclear genome of Selaginellaceae with respect to Isoetaceae. MP phylogenetic tree identified Isoetes subinermis, Isoetes durieui and Salvia microphylla as separate group among the studied taxa due to high sequence variation within these species through the time of evolution. Our result interpreted the polyphyletic origin of ferns and provides valuable information regarding the lycophytes and their fern allies.     

Molecular phylogenetics in 2D: ITS2 rRNA evolution and sequence-structure barcode from Veneridae to Bivalvia

In this study, we analyzed the nuclear ITS2 rRNA primary sequence and secondary structure in Veneridae and comparatively with 20 Bivalvia taxa to test the phylogenetic resolution of this marker and its suitability for molecular diagnosis at different taxonomic levels. Maximum likelihood and Bayesian trees based on primary sequences were congruent with (profile-) neighbor-joining trees based on a combined model of sequence-structure evolution. ITS2 showed higher resolution below the subfamily level, providing a phylogenetic signal comparable to (mitochondrial/nuclear) gene fragments 2-5 times longer. Structural elements of the ITS2 folding, such as specific mismatch pairing and compensatory base changes, provided further support for the monophyly of some groups and for their phylogenetic relationships. Veneridae ITS2 folding is structured in six domains (DI-VI) and shows five striking sequence-structure features. Two of them, the Basal and Apical STEMs, are common to Bivalvia, while the presence of both the Branched STEM and the Y/R stretches occurs in five superfamilies of the two Heterodonta orders Myoida and Veneroida, thus questioning their reciprocal monophyly. Our results validated the ITS2 as a suitable marker for venerids phylogenetics and taxonomy, and underlined the significance of including secondary structure information for both applications at several systematic levels within bivalves.     

The considerable genome size variation of Hordeum species (poaceae) is linked to phylogeny, life form, ecology, and speciation rates

Genome size variation in plants is thought to be correlatedwith cytological, physiological, or ecological characters. However,conclusions drawn in several studies were often contradictory.To analyze nuclear genome size evolution in a phylogenetic framework,DNA contents of 134 accessions, representing all but one speciesof the barley genus Hordeum L., were measured by flow cytometry.The 2C DNA contents were in a range from 6.85 to 10.67 pg indiploids (2n = 14) and reached up to 29.85 pg in hexaploid species(2n = 42). The smallest genomes were found in taxa from theNew World, which became secondarily annual, whereas the largestdiploid genomes occur in Eurasian annuals. Genome sizes of polyploidtaxa equaled mostly the added sizes of their proposed progenitorsor were slightly (1% to 5%) smaller. The analysis of ancestralgenome sizes on the base of the phylogeny of the genus revealedlineages with decreasing and with increasing genome sizes. Correlationsof intraspecific genome size variation with the length of vegetationperiod were found in H. marinum populations from Western Europebut were not significant within two species from South America.On a higher taxonomical level (i.e., for species groups or theentire genus), environmental correlations were absent. Thiscould mostly be attributed to the superimposition of life-formchanges and phylogenetic constraints, which conceal ecogeographicalcorrelations.     

Unexpected high divergence in nrDNA ITS and extensive parallelism in floral morphology of Pedicularis (Orobanchaceae)

Internal transcribed spacer (ITS) region sequences of nuclear ribosomal DNA (nrDNA) were used in the phylogenetic reconstruction of Pedicularis, a genus with strong adaptive radiation. Forty-two species representing 12 greges of the genus were all, except P. resupinata, sampled from the Hengduan Mountain region, China. A high level of ITS sequence variation was found among the species distributed in such a small geographical area, which had been rarely reported in other groups. The great ITS divergence in Pedicularis could be explained by a relatively ancient origin and diversification of the genus followed by migration of different floristic components into the Hengduan Mountains, or accelerated rates of molecular evolution in parasitic lineages of Orobanchaceae. In the present ITS phylogeny, almost all the main clades are not consistent with the high hierarchical taxa within Pedicularis, which implies that significant parallel evolution occurred in floral morphology of the genus, and that undue attention has been paid to corolla characters in the intragenus classifications.We thank the two anonymous reviewers for valuable comments and suggestions; Professor Guo You-Hao, Drs. Tan Dun-Yan, Luo Yi-Bo, Bao Ying, Luo Yan, Mr. Mao Jian-Feng and Mr. Qin Wei for their great help in the field collection; Drs. Song Bao-Hua, Wei Xiao-Xin and Miss Sun Ying-Xue for their help in the laboratory work. This study was supported by State Key Basic Research and Development Plan (Grant No. G2000046804) and the National Natural Science Foundation of China (Grant No. 30121003).     

樟科黄肉楠属是一个复系类群--基于nrDNA ITS和ETS序列分析

应用nrDNA ITS和ETS序列探讨了樟科Lauraceae黄肉楠属Actinodaphne的系统演化关系。对得到的3个序列矩阵（ITS、ETS和ITS／ETS）,采用MP（maximum parsimony）,ML（maximum likelihood）和Bayesian33分析方法进行了系统发育分析。结果显示,本文选的黄肉楠属Actinodaphne物种与所选的月棒族中的外类群靠近并混和在一起,进一步证实了本属为一个复系类群。结合对传统的形态学性状的重新认识,认为花序类型特征可能是重新界定黄肉楠属的最重要的性状,具有相同化序类型的物种可能具有相同的起源。然而,由于取样数量相对较少以及对矩阵的中．独分析存在一定的差异,还需更详细的研究来验证本文对黄肉楠属系统演化关系的假设,并进一步更精确地重建本属的系统发育关系。     

Phylogenetic inference of the genus Bupleurum (Apiaceae) in Hengduan Mountains based on chromosome counts and nuclear ribosomal DNA ITS sequences

pleurum of Apiaceae in China. This paper reports chromosome numbers of six species and two varieties of Bupleurum, and for four species and two varieties their chromosome numbers are reported for the first time. The phylogeny of Bupleurum was investigated based on the ITS region of the nuclear ribosomal DNA (nrDNA) of 14 taxa from the Hengduan Mountains, 3 taxa from the North China (Hebei and Heilongjiang), and 16 taxa from Africa and the Mediterranean region. Varia-tions in chromosome numbers and the ITS sequences were used to infer phylogenetic relationships between Bupleurum species in Hengduan Mountains. The results showed that the Hengduan Mountains might represent one of the frequency and diversity centers for Bupleurum. The ancestors of Bupleurum species in the Hengduan Mountains may be related to the woody B. fruticosum in North Africa, or the species in the western Mediterranean region. It is postulated that the ancestral population migrated into Hengduan Mountains through the Middle East and the Caucasus. Furthermore, the neo-endemic B. mundtii in South Africa appeared to be a close relative of the species in the Hengduan Mountains. In the trend of basic chromosome number evolution, x = 8 should be regarded as the ancestral basic number, while x = 6, 7 as the derived ones. The Bupleurum species in the Hengduan Moun-tains have been undergoing changes in the basic chromosome numbers or the ploidy level. The ITS phylogenetic tree showed that the Chinese species were divided into two clades: one with the basic chromosome number x = 8, and the other with x = 6, 7. The results rejected the previous infrageneric classification of Bupleurum in China. We further suggested to raise B. marginatum var. stenophyllum to species rank based on the combined evidence from morphology, karyology, pollen morphology, and the ITS phylogenetic tree.     

Coding of intraspecific nucleotide polymorphisms: A tool to resolve reticulate evolutionary relationships in the ITS of beech trees (Fagus L., Fagaceae)

The internal transcribed spacers ITS1 and ITS2 of the nuclear ribosomal DNA (rDNA) have recently been found to display remarkable intraspecific polymorphism, a feature suggested as limiting their value for phylogenetic reconstructions. A comparative study of oligonucleotide motives and intraindividual nucleotide variability across all species of the tree genus Fagus (beech) shows, however, that this intraspecific ITS polymorphism follows a particular pattern, which can be used to detect reticulation and ancient polymorphism within the genus. Coding ITS polymorphisms as phylogenetically informative characters, moreover, resulted in better‐resolved phylogenies than traditional ‘base‐per‐base’ maximum parsimony and maximum likelihood analyses.     

基于染色体计数和ITS序列初步探讨横断山区柴胡属植物(伞形科)的系统发育

横断山区是中国柴胡属Bupleurum植物的分布中心。本文对横断山区6个种2变种进行了染色体记数报道,其中4个种2变种是首次报道。对横断山区的10个种4个变种、中国北方（河北和黑龙江）的3个种的nrDNA ITS进行测序,同时从GenBank里面下载同属的来自非洲和地中海西部的16个nrDNA ITS序列数据,结合染色体数目变化结果,初步探讨了横断山区柴胡属植物的系统发育。结果表明横断山区可能是现代柴胡属植物的频度中心和多样分布中心之一。它们的祖先种可能是非洲北部的木本柴胡属植物B.fruticosum,或者是地中海西部的柴胡属植物,推测是通过中东和高加索扩散而形成的,其中与非洲南部特有种B.mundtii的亲缘关系也较近;染色体基数演化趋势是：8是较原始基数,6和7是次生基数,其染色体异基数变异和多倍化可能是物种形成、进化以及向外扩散的主要方式;在ITS系统发育树中,中国柴胡属植物染色体基数为8的种类聚为一支,染色体基数为6和7的种类聚为了一支,不支持舒璞等（1998）关于中国柴胡属的属下分类系统。结合已有的形态学、细胞学、孢粉学证据和ITS系统发育树,建议窄竹叶柴胡B.marginatum var.stenophyllum独立成种。     

Multiple hybrid formation in natural populations: concerted evolution of the internal transcribed spacer of nuclear ribosomal DNA (ITS) in North American Arabis divaricarpa (Brassicaceae)

DNA sequence variation of the internal transcribed spacer (ITS) region of nuclear ribosomal DNA from Arabis holboellii, A. drummondii, and its putative hybrid A. divaricarpa was analyzed to study hybrid speciation in a species system geographically covering nearly the entire North American continent. Based on molecular systematics the investigated species are better combined under the genus Boechera. Multiple intraindividual ITS copies were detected in numerous accessions of A. divaricarpa, and, to a minor extent, in the parental taxa. Comparative phylogenetic analysis demonstrates that reticulate evolution is common. Consequently, concerted evolution of ITS regions resulted in different types of ITS fragments not only in hybrid populations but also in one of the parental taxa, A. holboellii. Hybrid formation often occurred independently at different sites and at different times, which is reflected by ITS copies resampling the original parental sequence variation in different ways. Some biogeographic structuring of genetic diversity is apparent and mirrors postglacial migration routes. Hybridization, reticulation, and apomixis are assumed to be the major forces driving speciation processes in this species complex. Analysis of conserved regions and secondary structures of the ITS region provided no evidence that, in this system, hybrid ITS evolution is predominantly driven in a particular direction. However, two regions in the ITS1 and ITS2, respectively, show higher mutation rates than expected from outgroup comparisons. Strong evidence for the occurrence of apomixis in A. holboellii and A. divaricarpa has come from pollen size measurements and estimations of pollen quality, which favor the hypothesis that A. drummondii served as paternal hybridization partner more frequently than A. holboellii.     

Evidence for the Polyphyly of Haworthia (Asphodelaceae Subfamily Alooideae; Asparagales) Inferred from Nucleotide Sequences of rbcL, matK, ITS1 and Genomic Fingerprinting with ISSR-PCR

Abstract: Four molecular markers have been studied to examine the phylogenetic position of the South African plant genus Haworthia Duval within the succulent Asphodelaceae. Sequence data of the chloroplast genes mat K and rbc L were compared to the nuclear markers ITS1 and ISSR (Inter Simple Sequence Repeat) analysis. Both lines of molecular data, chloroplast and nuclear DNA, indicate that Haworthia is polyphyletic, forming two distinct clades. Most taxa previously combined as Haworthia subgenus Haworthia branch off early in the alooid chloroplast trees forming a strongly monophyletic group, whereas subgenus Hexangulares forms a polyphyletic assemblage comprising other alooid genera. The nuclear markers ITS1 and ISSR fingerprinting support the two groups as distinctly different, therefore confirming the division seen in chloroplast DNA. The practical implication is that the generic concept of Haworthia may have to be restricted to H. subgenus Haworthia or alternatively, that the groups of Haworthia be treated as infrageneric taxa within a broadened (Linnaean) concept of Aloe.     

ITS假基因对栎属系统学研究的影响及其对分子系统学研究的启示

核糖体rDNA ITS是被子植物系统发育研究中应用最广泛的分子标记之一。以前人们认为同一物种中的ITS序列因致同进化而使不同拷贝高度一致,在分子系统学研究中常以ITS1-5.8S-ITS2序列作为构建系统进化树的基础。近年来,在对一些被子植物的研究中发现这段序列在同一物种中具有多态性,有些拷贝中的5．8S区不具编码功能,人们把含有不具编码功能5．8S区的ITS1-5．8S-ITS2序列定义为ITS假基因序列,它对同源基因致同进化的假设形成了新的挑战。在诸多应用ITS序列重建系统进化关系的研究中,栎属系统学研究因ITS假基因的发现而倍受关注。本文以栎属为例回顾了ITS假基因的发现过程,分析了其对该属系统学研究的影响,为分子生物学在植物系统进化研究中的应用提供一些新的参考。     

Sequence variation in ITS spacers and 5.8S rDNA and relationship of E, St, P, Ns, Xm, and H genomes in the genera of Agropyron, Elytrigia, Leymus, Pascopyrum, Psathyrostachys, and Hordeum

Understanding species evolution and improvement requires information of their genome origin and differentiation. Among the species in the family Gramineae, genome identities of Agropyron-Elytrigia-Leymus group are still ambiguous. In order to delineate the genome relationship, nucleotide sequence analysis in the rDNA ITS regions was carried out among the species in the genera Elytrigia, Agropyron, Psathyrostachys, Leymus, and Psacopyrum containing E, St, P, Ns, and Xm genomes. The ITS-1 and ITS-2 showed a narrow range of variation in length except for the presence of a pentanucleotide, TGGGG, in/del in some haplotypes, whereas higher numbers of nucleotide substitutions were observed in most genera. There were 187 variable sites in the ITS-1, 5.8S, and ITS-2 regions, in which a few genome specific mutations were observed. While the level of variation was similar between ITS-1 and ITS-2, the rate of transition mutation versus transversion mutations was different among the ITS-1, 5.8S, and ITS-2 segments. GC contents of the ITS regions ranged between 55–65% between genomes and the haplotypes of P and H genomes were slightly higher than others. In phylogenetic analysis, the ITS haplotypes were classified into two groups; one containing H, Ns, NsXm genomes, and another containing P, St, and E genomes, which are congruous to the genome affinities from other studies. Among the four genomes in Pascopyrum smithii (2n=8x=56, StStNsNsHHXmXm), the haplotypes of H and St genomes were identified with the reference diploid species, but the haplotypes having Ns and Xm genomes were not found in the present analysis.     

Phylogeny and evolutionary history of the blister beetles (Coleoptera, Meloidae)

Meloid beetles are well characterised by both morphological and biological features. Previous phylogenetic hypotheses based on morphological characters assumed the repeated parallel evolution of complex biological novelties. In this work relationships among several taxa of the four subfamilies and almost all tribes representing meloid diversity are examined by using mitochondrial (16S) and nuclear (ITS2) DNA sequences, in 25 genera (using Anthicidae as outgroup). Secondary structure of 16S and ITS2 rRNAs were modelled. ITS2 structure represents a synapomorphic condition for the family and informative characters at the tribal level. Phylogenetic hypotheses based on separate and combined analysis of the 16S and ITS2 rDNA sequences, and morpho-biological characters were tested, and compared with previous morphological classifications. Molecular dating allowed an outline of the main steps of the evolutionary history of Meloidae, which evolved during Early Cretaceous and then radiated considerably with the adoption of hypermetaboly and parasitic behaviour, and with repeated, parallel evolution of larval phoresy on its hosts.     

基于核rDNA的ITS序列在种子植物系统发育研究中的应用

种子植物核rDNA是高度重复的串联序列,由于同步进化的力量．大多数物种中这些重复单位间已发生纯合或接近纯合。5．8S rDNA把核rDNA的内转录间隔区分为ITS1和ITS2两部分．在被子植物中ITS1的长度为165～298bp,ITS2的长度为177～266bp,而在裸子植物中ITS片段较长。且其长度变化主要由ITS1的长度变异所致。可对这两个片段PCR产物进行直接测序或克隆测序。由于ITS序列变异较快．能够提供较丰富的变异位点和信息位点,已成为被子植物较低分类阶元的系统发育和分类研究中的重要分子标记,为探讨多倍体复合体网状进化关系,异源多倍体的起源提供了重要的系统学信息．但它一般不适合科以上水平的系统学研究。裸子植物中ITS片段较长,重复序列间的纯合程度不同,测序比较困难．因此对探讨裸子植物系统发育和分类受到了一定的限制,但近年来有所发展。     

The evolution of CpNpG methylation in plants

Summary CpNpG and CpG methylation was surveyed in a range of vascular and nonvascular plants to determine firstly when CpNpG methylation evolved and secondly whether the two methylation systems found in higher plants were likely to be under common or separate control. Although both systems exist in a wide range of vascular plant taxa, the nonvascular plant taxa appear to contain only CpNpG methylation and this in only very limited amounts. The data suggest that both systems may have evolved at the same time and that speciation involved loss of one or the other methylation system or the evolution of differentiation stage-specific control systems.     

ITS1 Sequences of Nuclear Ribosomal DNA in Wild Rices and Cultivated Rices of China and Their Phylogenetic Implications

he first internal transcribed spacer (ITS1) of nuclear ribosomal DNA of three wild rice species and two subspecies of cultivated rice, which are distributed in China, was amplified using PCR technique and sequenced with automated fluorescent sequencing. The sequences of ITS1 ranged from 193 bp to 218 bp in size and G/C content varied from 69.3%to 72.7%. In pairwise comparison among the five taxa, sequence site divergence ranged from 1.5 % to 10.6%. Phylogenetic analysis of ITS1 sequences using Wagner parsimony generated a single well-resolved tree, which revealed that Oryza rufipogon was much more closely related to cultivated rice species than to the other two wild species. Oryza granulata was less closely related to either cultivated rice species or the other two wild species, and might be a unique and isolated taxon in the genus Oryza. The phylogenetic relationships of the three wild rice species and two cultivated rice subspecies inferred from ITS1 sequences is highly concordant with those based on the molecular evidence from isozyme, chloroplast DNA (cpDNA), mitochondrial DNA (mtDNA) and nuclear DNA (nDNA) of the genus Oryza.