分子进化生物学中序列分析方法的新进展

简要介绍了分子进化生物学中序列分析方法的最新进展,特别强调了似然比检验和贝叶斯推论在分子进化和系统发育假说检验中的重要性,并介绍了新方法的一些成功应用,同时还给出了一些重要的信息资源。     

序列消除与异源多倍体植物基因组的进化

经杂交后多倍化形成的异源多倍体植物,被认为在其形成的早期阶段经历了DNA序列消除过程。发生消除的序列既涉及到高拷贝的序列也有低拷贝的序列,而且大多数情况下倾向于消除来自其中一个亲本的序列。序列消除的模式因基因组组成和物种的不同而有差异,并且可能受到细胞质的影响。尽管序列消除的分子机制还不是很清楚,但很多证据已表明非同源染色体之间的互作不是主要的原因。目前认为,序列消除增加了非同源染色体之间的差异,为多倍化后在减数分裂过程中快速恢复二倍化的染色体配对模式提供了物质基础,这样更有利于多倍体在自然界快速稳定。     

DNA序列进化过程中核苷酸替代的非独立性研究

本文评述了DNA序列间核苷酸替代数的估计方法,并通过对七个物种中组蛋白基因的比较对DNA进化的模型进行了考察。发现H2A基因第三位点上的碱基组成在物种间变异很大,并且跟H2A基因第一位点、H4基因第一、三位点及H2A上游,下游序列中的碱基组成有强正相关,提示DNA序列进化过程中存在着物种特异的区域性约束力。可能的原因是高等真核生物中GC含量升高,或者是染色体重组使这些同源序列位于不同的等质区段,从而受到不同的选择突变压。密码内各位点上核苷酸替代的相关性分析表明不同位点的替代是非独立的,其原因可能是一次替代事件引起多个位点的变化。文中讨论了这些结果对进化树推断的意义。     

分子进化研究中系统发生树的重建

在现代分子进化研究中,根据现有生物基因或物种多样性来重建生物的进化史是一个非常重要的问题。一个可靠的系统发生的推断,将揭示出有关生物进化过程的顺序,有助于我们了解生物进化的历史和进化机制。本文简要介绍了系统发生树推断的几个重要问题：建树方法、数据转换、树的可靠性及目前使用较多的几种分析软件。     

《计算分子进化》（复旦大学进化生物学丛书）

进化生物学在近二十年经历了一个快速发展和变革的时期,是当今生命科学领域发展最为迅速的分支学科之一。后基因组时代的生物进化研究不仅在研究思路和研究方法上突破了传统的研究模式,在揭示生物起源和进化机制方面向前迈出了一大步,而且极大地拓展了研究领域,向具有广泛的社会实用性的方向转变,在揭示人类重大遗传疾病的分子基础、传染性疾病爆发与病原生物进化变异的关系、以及生物对环境变化的响应和适应机制方面显示出巨大的潜力。     

小麦A，B和D基因组同源DNA序列在进化过程中的演变研究

选取已定位的大麦1H染色体的STS标记NWG913为引物,在普通小麦（Tritium aestivum L.）及其4个可能的起源种乌拉尔图小麦（T.urartu T.)、栽培一粒小麦（T.monococcum.L)栽培二粒小麦（T.dicoccum S.）、方穗山羊草（Ae.squarrosa L.)上特异性扩增。扩增产物克隆测序后对其进行序列分析,由序列差异的程度来确定这几个物种之间的亲缘关系。实验结果表明,普通小麦（Tritium aestivum L.)的A基因组此段序列与乌拉尔图小麦（T.urartu T.)、栽培一粒小麦（T.monococcum L.)、栽培二粒小麦（T.dicoccum S.)A基因组此段序列完全相同;普通小麦的D基因组此段序列与方穗山羊草（Ae.squarrosa L.)也完全相同;普通小麦的B基因组此段序列和栽培二粒小麦B基因组此段序列有0．61％的差异。研究结果一方面对现有的普通小麦A、B、D基因组起源和进化理论给予了分子水平上的证明,同时也揭示了同一物种不同的基因组化进化速度存在差异。     

果蝇中顺式调控序列的进化

顺式调控序列(cis-regularory sequences)是与基因表达调控相关、能够被调控因子特异性识别和结合的非编码DNA序列。顺式调控序列可以通过增减所含转录因子结合位点的数目,重构转录调控网络,以精准调控基因的时空表达模式,从而调控动物的生理和形态表型。顺式调控假说认为顺式调控序列进化是自然界丰富的动物表型进化的主要遗传机制。本文首先简述了顺式调控序列的结构和功能,然后重点讨论了近年来顺式调控序列进化调控果蝇表型进化如刚毛表型进化、色素沉积表型进化和胚胎发育方面的研究进展,阐释了顺式调控序列进化是动物表型进化的主要遗传机制之一。最后本文展望了顺式调控序列未来研究方向,例如应用功能基因组研究、开展ENCODE计划等,为进化发育生物学中顺式调控序列的研究提供了参考。     

银色裂腹鱼线粒体基因组序列特征与系统进化分析

银色裂腹鱼(Schizothorax argentatus)在我国仅分布于新疆地区的伊犁河流域,是我国裂腹鱼类中珍稀濒危品种之一,具有较高的科研和经济价值。本研究采用高通量测序技术获得了银色裂腹鱼长度为16580 bp的线粒体基因组全序列,其基因组成和排列顺序均与典型的脊椎动物相似,共有13个蛋白质编码基因、22个tRNA基因、2个rRNA基因和1个非编码区(D-loop)。碱基组成分别为A(30.25%)、G(17.28%)、C(27.20%)和T(25.27%),呈现明显的AT偏好性和反G偏倚。tRNA基因中仅tRNA-Ser(GCU)因缺少二氢尿嘧啶茎而无法形成典型的三叶草结构。ND6基因的AT-skew和GC-skew值波动最大,揭示该基因经历的选择和突变压力可能与其他基因不同。银色裂腹鱼线粒体控制区包含了3个不同的结构域:终止序列区(ETAS)、中央保守区(CSB-F、CSB-E、CSB-D和CSB-B)和保守序列区(CSB1、CSB2和CSB3),且在CSB3下游约50 bp处识别到鲤形目(Cypriniformes)鱼类中普遍存在的保守序列片段TT(AT)nGTG。基于28种裂腹鱼属鱼类线粒体基因组全序列构建的系统发育关系表明银色裂腹鱼分化时间较早,与其他类群亲缘关系较远,这可能与其所生活的水域地理位置和水文环境有密切关系。     

浙江省汉坦病毒基因分子进化分析

为了分析浙江省汉坦病毒分子流行病学的特点,本文应用分子生物学软件对1981～2007年从浙江省不同地区不同宿主分离的12株汉坦病毒的S、M基因序列进行分子进化分析,结果显示:浙江省属汉滩(HTN)型和汉城(SEO)型的混合型疫区,病毒的基因差异和亲缘远近关系主要表现在地区性,而与病毒分离的年代与宿主关系并不大,表现出明显的地理聚集现象,该现象在HTNV中的表现尤为明显,同一地区分离的病毒表现为较高的基因同源性,系统进化树上分布于同一或临近分支。另外发现分离自浙江省建德地区的Gou3和ZJ5株在SEOV进化枝中构成一独立分支,而且与国内外SEOV其他毒株的基因差异均较大,在浙江省建德地区存在着SOEV的特殊亚型病毒。     

木聚糖酶分子进化的研究进展

木聚糖的降解需要多种水解酶的协同作用,其中βD1,4内切木聚糖酶是最关键的水解酶之一。同一家族木聚糖酶氨基酸序列间有较高的同源性和较近的亲缘关系,这标准通常用于酶的家族归类。不同来源的同种木聚糖酶在相同位置上的氨基酸残基起源于共同祖先或者具有相似的生物学功能,而在进化过程中,对酶分子结构、催化起重要作用的氨基酸残基往往高度保守。综述了木聚糖酶分子进化的研究进展及其应用前景 。     

Hepatitis C Virus Evolutionary Patterns Studied Through Analysis of Full-Genome Sequences

The evolutionary patterns of hepatitis C virus (HCV), including the best-fitting nucleotide substitution model and the molecular clock hypothesis, were investigated by analyzing full-genome sequences available in the HCV database. The likelihood ratio test allowed us to discriminate among different evolutionary hypotheses. The phylogeny of the six major HCV types was accurately inferred, and the final tree was rooted by reconstructing the hypothetical HCV common ancestor with the maximum likelihood method. The presence of phylogenetic noise and the relative nucleotide substitution rates in the different HCV genes were also examined. These results offer a general guideline for the future of HCV phylogenetic analysis and also provide important insights on HCV origin and evolution. Received: 13 January 2001 / Accepted: 21 June 2001     

How do insect nuclear and mitochondrial gene substitution patterns differ? Insights from Bayesian analyses of combined datasets

We analyzed 12 combined mitochondrial and nuclear gene datasets in seven orders of insects using both equal weights parsimony (to evaluate phylogenetic utility) and Bayesian methods (to investigate substitution patterns). For the Bayesian analyses we used relatively complex models (e.g., general time reversible models with rate variation) that allowed us to quantitatively compare relative rates among genes and codon positions, patterns of rate variation among genes, and substitution patterns within genes. Our analyses indicate that nuclear and mitochondrial genes differ in a number of important ways, some of which are correlated with phylogenetic utility. First and most obviously, nuclear genes generally evolve more slowly than mitochondrial genes (except in one case), making them better markers for deep divergences. Second, nuclear genes showed universally high values of CI and (generally) contribute more to overall tree resolution than mitochondrial genes (as measured by partitioned Bremer support). Third, nuclear genes show more homogeneous patterns of among-site rate variation (higher values of alpha than mitochondrial genes). Finally, nuclear genes show more symmetrical transformation rate matrices than mitochondrial genes. The combination of low values of alpha and highly asymmetrical transformation rate matrices may explain the overall poor performance of mitochondrial genes when compared to nuclear genes in the same analysis. Our analyses indicate that some parameters are highly correlated. For example, A/T bias was positively and significantly associated with relative rate and CI was positively and significantly associated with alpha (the shape of the gamma distribution). These results provide important insights into the substitution patterns that might characterized high quality genes for phylogenetic analysis: high values of alpha, unbiased base composition, and symmetrical transformation rate matrices. We argue that insect molecular systematists should increasingly focus on nuclear rather than mitochondrial gene datasets because nuclear genes do not suffer from the same substitutional biases that characterize mitochondrial genes.     

A Primer to Molecular Phylogenetic Analysis in Plants

Reconstructing a tree of life by inferring evolutionary history is an important focus of evolutionary biology. Phylogenetic reconstructions also provide useful information for a range of scientific disciplines such as botany, zoology, phylogeography, archaeology and biological anthropology. Until the development of protein and DNA sequencing techniques in the 1960s and 1970s, phylogenetic reconstructions were based on fossil records and comparative morphological/physiological analyses. Since then, progress in molecular phylogenetics has compensated for some of the shortcomings of phenotype-based comparisons. Comparisons at the molecular level increase the accuracy of phylogenetic inference because there is no environmental influence on DNA/peptide sequences and evaluation of sequence similarity is not subjective. While the number of morphological/physiological characters that are sufficiently conserved for phylogenetic inference is limited, molecular data provide a large number of datapoints and enable comparisons from diverse taxa. Over the last 20 years, developments in molecular phylogenetics have greatly contributed to our understanding of plant evolutionary relationships. Regions in the plant nuclear and organellar genomes that are optimal for phylogenetic inference have been determined and recent advances in DNA sequencing techniques have enabled comparisons at the whole genome level. Sequences from the nuclear and organellar genomes of thousands of plant species are readily available in public databases, enabling researchers without access to molecular biology tools to investigate phylogenetic relationships by sequence comparisons using the appropriate nucleotide substitution models and tree building algorithms. In the present review, the statistical models and algorithms used to reconstruct phylogenetic trees are introduced and advances in the exploration and utilization of plant genomes for molecular phylogenetic analyses are discussed.     

Accuracy and Power of the Likelihood Ratio Test for Comparing Evolutionary Rates Among Genes

Sequences for multiple protein-coding genes are now commonly available from several, often closely related species. These data sets offer intriguing opportunities to test hypotheses regarding whether different types of genes evolve under different selective pressures. Although maximum likelihood (ML) models of codon substitution that are suitable for such analyses have been developed, little is known about the statistical properties of these tests. We use a previously developed fixed-sites model and computer simulations to examine the accuracy and power of the likelihood ratio test (LRT) in comparing the nonsynonymous-to-synonymous substitution rate ratio (=dN/dS) between two genes. Our results show that the LRT applied to fixed-sites models may be inaccurate in some cases when setting significance thresholds using a ² approximation. Instead, we use a parametric bootstrap to describe the distribution of the LRT statistic for fixed-sites models and examine the power of the test as a function of sampling variables and properties of the genes under study. We find that the power of the test is high (>80%) even when sampling few taxa (e.g., six species) if sequences are sufficiently diverged and the test is largely unaffected by the tree topology used to simulate data. Our simulations show fixed-sites models are suitable for comparing substitution parameters among genes evolving under even strong evolutionary constraint ( 0.05), although relative rate differences of 25% or less may be difficult to detect.Reviewing Editor: Dr. Rosmus Nielsen     

An Assessment of the Systematics of Arvicanthine Rodents Using Mitochondrial DNA Sequences: Evolutionary and Biogeographical Implications

Mitochondrial DNA sequences of cytochrome b (1140-bp), 12S (375-bp) and 16S (475-bp) ribosomal RNA gene fragments were used to investigate the phylogenetic relationships of a group of African rodents referred as the arvicanthines (Family Muridae, Subfamily Murinae). A total of 49 specimens including all seven genera and 15 of the 24 arvicanthine species currently recognized as well as outgroups from the subfamily Acomyinae, Arvicolinae, Gerbillinae, Murinae and Otomyinae were examined. Our molecular data support the monophyly of the African arvicanthine genera and their partition into three distinct lineages: one composed of Arvicanthis, Mylomys and Pelomys, one composed of Desmomys and Rhabdomys, and one represented by Lemniscomys. The Indian arvicanthine Golunda is external to this clade and is part of a larger clade, together with the African arvicanthines and other African Murinae such as Aethomys, Dasymys, Grammomys, and Hybomys, for which we propose the use of the tribal name Arvicanthini. The basal relationships within this set of species are poorly resolved, suggesting the possibility of a rapid radiation. Calibration based on the fossil record suggests that this radiative event would have taken place at about 8.0 Mya (Million years ago). The identification of the Otomyinae as the sister-taxon to Arvicanthini implies that the former are true murines and should therefore be given only tribal rank within the Murinae.     

Towards the phylogeny of the Curculionoidea (Coleoptera): Reconstructions from mitochondrial and nuclear ribosomal DNA sequences

With about 60,000 described species, Curculionoidea represent the most species-rich superfamily in the animal kingdom. The immense diversity apparently creates difficulties in the reconstruction of the phylogenetic relationships. Independent morphological studies have led to very different classifications. This study is based on molecular data from two independent molecular sources, the 16S and 18S rDNA. Sensitivity analyses were conducted for the sequence alignment (gap costs were varied) as well as the phylogenetic reconstruction algorithms and some of their parameters. The higher-level relationships reconstructed within Curculionoidea are sensitive to alignment and reconstruction method. Nemonychidae or Oxycorynidae+Belidae were found to be sister to all remaining Curculionoidea in many analyses. The 16S rDNA sequence data (obtained from 157 species) corroborate many tribes and genera as monophyletic. It is observed that the phylogenetic reconstruction of genera with specific genetic features such as polyploidy and parthenogenetic reproduction is difficult in weevils. The curculionid subfamily Lixinae appears monophyletic. A new monophylum consisting of Entiminae, Hyperinae, Cyclominae, Myllorhinus plus possibly the Cossoninae is distinguished and we call it Entiminae s.l. For most other subfamilies and families homoplasy concealed the phylogenetic signal (due to saturation of the 16S sequences), or the species sampling was insufficient, although our sampling scheme was rather broad. We observed that although data from one source can easily be misleading (16S) or hardly informative (18S), the combination of the two independent data sets can result in useful information for such a speciose group of organisms. Our study represents the most thorough analysis of molecular sequence data of the Curculionoidea to date and although the phylogenetic results appear less stable than expected, they reflect the information content of these sequence data realistically and thus contribute to the total knowledge about the phylogeny of the Curculionoidea.     

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。     

Molecular phylogenies of plastid origins and algal evolution

Summary An overview of recent molecular analyses regarding origins of plastids in algal lineages is presented. Since different phylogenetic analyses can yield contradictory views of algal plastid origins, we have examined the effect of two distance measurement methods and two distance matrix tree-building methods upon topologies for the ribulose-1,5-bisphosphate carboxylase/oxygenase large subunit nucleotide sequence data set. These results are contrasted to those from bootstrap parsimony analysis of nucleotide sequence data subsets. It is shown that the phylogenetic information contained within nucleotide sequences for the chloroplast-encoded gene for the large subunit of ribulose-1,5-bisphosphate carboxylase/oxygenase, integral to photosynthesis, indicates an independent origin for this plastid gene in different plant taxa. This finding is contrasted to contrary results derived from 16S rRNA sequences. Possible explanations for discrepancies observed for these two different molecules are put forth. Other molecular sequence data which address questions of early plant evolution and the eubacterial origins of algal organelles are discussed.Offprint requests to: W. Martin     

重要观赏兰科植物的分子生物学研究进展

兰科植物是开花植物中最大的家族之一,分子标记技术应用于兰科植物的分类鉴定和品种鉴别,为兰花的分类提供了分子水平的证据,也为兰花保护策略和措施的制定提供了理论基础.兰科植物表现有高度特异的形态、结构和生理特性,是研究花着色机理和子房发育的理想对象.兰花离体培养开花系统的建立,可以用来探明兰花从营养生长向生殖生长的转变机制,是研究花的分化和发育的理想材料.兰花具有特异的查尔酮合成酶(CHS)基因和二氢叶酸还原酶(DFR)基因等控制花色素的合成,DOHI基因控制石斛兰花芽的形成和提早开花,PHAL039基因和ACC合成酶基因在蝴蝶兰授粉后的子房发育中起着重要的调控作用,这些特异基因的分离和克隆为兰花花的分化、发育及着色机制提供了分子基础.蝴蝶兰属、大花蕙兰(Cymbidium hybrdium)、石斛兰属、文心兰属、五唇兰属和万代兰属等兰科植物都有转基因的研究报道,主要以原球茎为材料采用基因枪或农杆菌法转化,部分研究获得了转化植株.