基于COXI基因的太平洋西岸文昌鱼地理种群分析

文昌鱼是进化发育研究的重要模式动物,目前实验材料均采自野外.因此,对其进行正确的物种鉴定和地理种群的遗传分化分析十分必要.该研究扩增并测定了COX 1基因部分序列,结合NCBI数据库中的COX 1序列信息,对太平洋西岸文昌鱼的种类和地理种群分化情况进行了分析.结果表明,马来文昌鱼(Branchiostoma malayanum)、白氏文昌鱼(B.belcheri)和日本文昌鱼(B.japonicum)这3个种之间的遗传差异很大,再次证实3个物种的有效性,同时提出应当审慎对待NCBI数据库中文昌鱼的种名标注;太平洋西岸文昌鱼属Branchiostoma的3种文昌鱼群体遗传多样性均处于较高水平,同一物种的不同地理种群间没有出现明显的遗传分化,反映了海洋动物的基因交流较容易,不同海域隔离较弱.     

偏文昌鱼属(Asymmetron)在中国海的发现和厦门文昌鱼的地理分布

原索动物门(Phylum Protochordata)中的头索功物纲(Classe Cephalochordata),现在一般公认的仅有一科两属,即文昌鱼科(Famille Branchiostomidae)的文昌鱼属(GenreBranchiostoma Costa)和偏交昌鱼属(Genre Asymmetron Andrews)。文昌鱼属的厦门文昌角[Branchiostoma belcheri(Gray)]早在1923年就在我国厦门刘五店大量发现,它和它的变种青岛文昌鱼(B.belcheri var.tsingtauense Tchang et Koo)广泛地分布在中国海的近     

厦门两种文昌鱼染色体的制备与观察

文吕鱼的进化地位十分重要,对其染色体的研究在进化和比较基因组学方面有重要意义.然而文昌鱼的染色体制备困难,使研究受到了限制.本文介绍了一种改良的义昌鱼胚胎细胞染色体标本制备方法,以及用文昌鱼成体再生细胞制备染色体,首次获得了文昌鱼体细胞中期染色体标本,并观察了厦门2种文昌鱼的染色体,其中白氏文吕鱼(Branchiostoma belcheri)二倍体2n=40,日本文昌鱼(B.japonicum)二倍体2n=36.再次从细胞分类学角度证实白氏文昌鱼和日本文昌鱼作为两个独立物种的分类地位.     

通过Cyt b基因同源序列比较评估厦门文昌鱼的分类学地位

白氏文昌鱼Branchiostomabelcheri (Gray)在我国和日本沿海均有分布 ,由于南、北方文昌鱼形态学上有一定差异 ,且二者间存在一些过渡类型 ,其分类地位问题仍有待进一步澄清。本文测定了厦门欧厝海域产的文昌鱼mtDNACytb基因序列 ,并与日本产的文昌鱼以及另外产于大西洋的两种文昌鱼Cytb基因序列比较。分子系统学分析结果表明 :厦门欧厝海域产的文昌鱼与日本产的文昌鱼平均遗传距离为 2 1 12 % ,达到了种间分化的水平 ;经过对已有文献和文昌鱼地理分布的综合分析 ,作者建议将原来的白氏文昌鱼青岛亚种B belcheritsingtauense提升为种 ,南、北方所产文昌鱼分别作为两个独立的种存在 ,即南方的B belcheri (Gray)和北方的B tsingtauenseTchangetKoo     

文昌鱼的实验室繁育及子二代获得

文昌鱼特殊的进化地位、简单的器官系统和终生透明的躯体等特征,使其很有希望成为一个新型实验室模式动物。要实现文昌鱼的模式动物化,实验室内全人工条件下繁殖是关键的第一步。为此,我们于2003年9月和2004年4月两次采集产于厦门海域的2种文昌鱼,开展实验室内养殖研究。经过3年多的持续实验室养殖,继2005年夏季于实验室内繁殖出子一代文昌鱼后,又在2006年夏季成功获得了这两种文昌鱼的子二代,初步实现文昌鱼在实验室内的全人工养殖。对子代文昌鱼养殖的初步观察发现,不同水温对生长发育速度有一定影响,提示有可能通过水温控制实现文昌鱼一年多次产卵的目的。目前这两种文昌鱼子二代幼体已完成变态,进入亚成体生长发育阶段,其体长分别已达14.6mm(日本文昌鱼Branchiostomajaponicum)和6.5mm(白氏文昌鱼B.belcheri)。     

文昌鱼AmphiRab23b基因的克隆、进化分析及表达模式

Hedgehog信号通路在胚胎发育过程中发挥着重要作用, 同时与多种肿瘤的发生密切相关。Rab23蛋白在Hedgehog信号通路中扮演着十分重要的角色。目前关于文昌鱼Rab23同源基因的研究仅局限于佛罗里达文昌鱼(Branchiostoma floridae)基因组中的注释。文章首次克隆了中国文昌鱼(Branchiostoma belcheri) Rab23b基因 (AmphiRab23b)cDNA全序列 , 对其演译的蛋白序列进行了序列比对、进化树分析以及基因时空表达分析。研究结果显示, 文昌鱼AmphiRab23b基因的 cDNA总长为2 062 bp (包括UTR区), 其中开放阅读框 (Open reading frame, ORF) 714 bp, 编码237个氨基酸; 虽然在进化树中不属于脊椎动物Rab23进化支, 但是AmphiRab23具有保守的Rab23_lke结构域, 暗示该基因在进化过程中可能在功能上是保守的。时空表达的研究结果进一步显示, AmphiRab23b基因在胚胎发育中的神经板和消化道中表达, 与其脊椎动物同源基因的表达模式相似, 这说明该基因可能对文昌鱼神经系统和消化道的发育有重要作用。     

文昌鱼—研究脊柱动物起源和进化的模式动物

长久以来,文昌鱼一直被认为和生活在约5亿年前的脊椎动物的直接祖先相似。由于文昌鱼在进化上的重要性,它在动物学研究史上发挥着关键作用,近100多年来,文昌鱼作为研究对象曾数次受到动物学界青睐或冷落,大约10年前,随着分子生物学技术应用于文昌鱼研究,又激发了动物学家对文昌鱼的研究兴趣,又一次出现在文昌鱼研究的高潮,并且一直持续至今,分子生物学研究结果表明,文昌鱼样生物可能是环节动物样动物和最早的脊椎动物之间的进化中间体,因此,文昌鱼在动物学研究史上好像绕了个大圈又回到了原处,在被忽视一段时间之后,又重新占据脊椎动物起源和进化研究中心舞台的位置,成为研究脊椎动物起源和进化的模式动物。     

厦门两种文昌鱼染色体的制备与观察(英文）

文昌鱼的进化地位十分重要,对其染色体的研究在进化和比较基因组学方面有重要意义。然而文昌鱼的染色体制备困难,使研究受到了限制。本文介绍了一种改良的文昌鱼胚胎细胞染色体标本制备方法,以及用文昌鱼成体再生细胞制备染色体,首次获得了文昌鱼体细胞中期染色体标本,并观察了厦门2种文昌鱼的染色体,其中白氏文昌鱼（Branchiostoma belcheri）二倍体2n=40,日本文昌鱼（B. japonicum）二倍体2n=36。再次从细胞分类学角度证实白氏文昌鱼和日本文昌鱼作为两个独立物种的分类地位。     

厦门文昌鱼早期胚胎发育的观察

文昌鱼的胚胎发育是脊索动物的典型范例。欧洲文昌鱼Branchiostoma lanceplatum Pallas的正常发育已由(1866)、Hatschek(1881)和Conklin(1932)等详细地描述过。厦门文昌鱼B.belcheri Gray的基本构造及发育过程和欧洲的相似。是动物学和胚胎学教学上及实验胚胎学研究上的理想材料,所以正常发育规律的研究有一定的意义。本文着重分析性腺成熟时期的外界条件及受精和早期发育的过程。     

白氏文昌鱼单个体基因组BAC文库的构建

隶属于头索动物亚门的文昌鱼是现存生物中最近似于脊椎动物亚门直接祖先的一个类群, 具有重要的进化地位, 是研究脊椎动物原始祖先的重要材料和模式动物。随着文昌鱼实验室连续繁殖的成功, 全基因组测序成为中国文昌鱼模式化急需完成的工作之一。文章从单条雄性白氏文昌鱼精巢组织中提取高质量的基因组DNA, 经EcoRⅠ限制性内切酶和EcoRⅠ甲基化酶酶切, 脉冲场电泳选择合适酶切DNA片段, 连接线性磷酸化的载体pCC1BAC, 转化大肠杆菌EPI300 E. coli, 构建了含有44 706个克隆的全基因组BAC( Bacterial artificial chromosome)文库, 该文库平均插入片段80 kb,具有9倍的基因组覆盖率, 基本能够满足功能基因等研究需要, 为中国文昌鱼全基因组测序打下基础。     

Characterization of Evolutionarily Conserved MicroRNAs in Amphioxus

Amphioxus is an extant species closest to the ancestry of vertebrates.Observation of microRNA(miRNA)distribution of amphioxus would lend some hints for evolutionary research of vertebrates.In this study,using the publicly available scaffold data of the Florida amphioxus(Branchiostoma floridae)genome,we screened and characterized homologs of miRNAs that had been identified in other species.In total,68 pieces of such homologs were obtained and classified into 33 families.Most of these miRNAs were distributed as clusters in genome.Inter-species comparison showed that many miRNAs,which had been thought as vertebrate-or mammal-specific before,were also present in amphioxus,while some miRNAs that had been considered as protostome-specific before also existed in amphioxus.Compared with ciona,amphioxus had an apparent miRNA gene expansion,but phylogenetic analysis showed that the duplicated miRNAs or clusters of amphioxus had a higher homology level than those duplicated ones in vertebrates.     

Exploring developmental, functional, and evolutionary aspects of amphioxus sensory cells

Amphioxus has neither elaborated brains nor definitive sensory organs, so that the two may have evolved in a mutually affecting manner and given rise to the forms seen in extant vertebrates. Clarifying the developmental and functional aspects of the amphioxus sensory system is thus pivotal for inferring the early evolution of vertebrates. Morphological studies have identified and classified amphioxus sensory cells; however, it is completely unknown whether the morphological classification makes sense in functional and evolutionary terms. Molecular markers, such as gene expression, are therefore indispensable for investigating the developmental and functional aspects of amphioxus sensory cells. This article reviews recent molecular studies on amphioxus sensory cells. Increasing evidence shows that the non-neural ectoderm of amphioxus can be subdivided into molecularly distinct subdomains by the combinatorial code of developmental cues involving the RA-dependent Hox code, suggesting that amphioxus epithelial sensory cells developed along positional information. This study focuses particularly on research involving the molecular phylogeny and expression of the seven-transmembrane, G protein-coupled receptor (GPCR) genes and discusses the usefulness of this information for characterizing the sensory cells of amphioxus.     

Evidence for stasis and not genetic piracy in developmental expression patterns of Branchiostoma lanceolatum and Branchiostoma floridae, two amphioxus species that have evolved independently over the course of 200 Myr

Cephalochordates, the most basal extant group in the phylum Chordata, are represented chiefly by about 20 species of the genus Branchiostoma, commonly called amphioxus or lancelets. In recent years, insights into the evolutionary origin of the vertebrates have been gained from molecular genetic studies during the development of three of these amphioxus species (Branchiostoma floridae in North America, Branchiostoma lanceolatum in Europe, and Branchiostoma belcheri in East Asia). In spite of an estimated divergence time of 100-200 Myr among these species, all three are remarkably similar morphologically, and students of amphioxus have tacitly assumed that such resemblances arise during ontogeny from nearly identical networks of developmental genes. We felt that this assumption needed to be reexamined because instances are known--even in comparisons of closely related species--where characters seeming homologous on the basis of morphology actually develop under the control of conspicuously divergent genetic programs (a phenomenon termed "genetic piracy"). In the present work, we tested the hypothesis that morphological similarities reflect strict conservation of developmentally important genes' expression patterns in order to assess whether the developmental genetics of different amphioxus species show evidence of genetic piracy. To these ends, we cloned 18 genes implicated in different developmental functions in B. lanceolatum and compared their gene expression patterns with the known expression patterns of their orthologous genes in B. floridae. We show that, for the most part, conservation of gene expression parallels that of morphology in these two species. We also identified some differences in gene expression, likely reflecting experimental sensitivity, with the exception of Pax1/9, which may result from true developmental specificities in each amphioxus species. Our results demonstrate that morphological conservation reflects stasis in developmental gene expression patterns and find no evidence for genetic piracy. Thus, different species of amphioxus appear to be very similar, not only morphologically, but also in the genetic programs directing the development of their structural features. Moreover, we provide the first catalogue of gene expression data for the European species, B. lanceolatum.     

EST and transcriptome analysis of cephalochordate amphioxus--past, present and future

The cephalochordates, commonly known as amphioxus or lancelets, are now considered the most basal chordate group, and the studies of these organisms therefore offer important insights into various levels of evolutionary biology. In the past two decades, the investigation of amphioxus developmental biology has provided key knowledge for understanding the basic patterning mechanisms of chordates. Comparative genome studies of vertebrates and amphioxus have uncovered clear evidence supporting the hypothesis of two-round whole-genome duplication thought to have occurred early in vertebrate evolution and have shed light on the evolution of morphological novelties in the complex vertebrate body plan. Complementary to the amphioxus genome-sequencing project, a large collection of expressed sequence tags (ESTs) has been generated for amphioxus in recent years; this valuable collection represents a rich resource for gene discovery, expression profiling and molecular developmental studies in the amphioxus model. Here, we review previous EST analyses and available cDNA resources in amphioxus and discuss their value for use in evolutionary and developmental studies. We also discuss the potential advantages of applying high-throughput, next-generation sequencing (NGS) technologies to the field of amphioxus research.     

Genomics reveal ancient forms of stanniocalcin in amphioxus and tunicate

Stanniocalcin (STC) is present throughout vertebrates, including humans, but a structure for STC has not been identified in animals that evolved before bony fish. The origin of this pleiotropic hormone known to regulate calcium is not clear. In the present study, we have cloned three stanniocalcins from two invertebrates, the tunicate Ciona intestinalis and the amphioxus Branchiostoma floridae. Both species are protochordates with the tunicates as the closest living relatives to vertebrates. Amphioxus are basal to both tunicates and vertebrates. The genes and predicted proteins of tunicate and amphioxus share several key structural features found in all previously described homologs. Both the invertebrate and vertebrate genes have four conserved exons. The predicted length of the single pro-STC in Ciona is 237 amino acids and the two pro-hormones in amphioxus are 207 and 210 residues, which is shorter than human pro-STCs at 247 and 302 residues due to expansion of the C-terminal region in vertebrate forms. The conserved pattern of 10 cysteines in all chordate STCs is crucial for identification as amphioxus and tunicate amino acids are only 14-23% identical with human STC1 and STC2. The 11th cysteine, which is the cysteine shown to form a homodimer in vertebrates, is present only in amphioxus STCa, but not in amphioxus STCb or tunicate STC, suggesting the latter two are monomers. The expression of stanniocalcin in Ciona is widespread as shown by RT-PCR and by quantitative PCR. The latter method shows that the highest amount of STC mRNA is in the heart with lower amounts in the neural complex, branchial basket, and endostyle. A widespread distribution is present also in mammals and fish for both STC1 and STC2. Stanniocalcin is a presumptive regulator of calcium in both Ciona and amphioxus, although the structure of a STC receptor remains to be identified in any organism. Our data suggest that amphioxus STCa is most similar to the common ancestor of vertebrate STCs because it has an 11th cysteine necessary for dimerization, an N-glycosylation motif, although not the canonical one in vertebrate STCs, and similar gene organization. Tunicate and amphioxus STCs are more similar in structure to vertebrate STC1 than to vertebrate STC2. The unique features of STC2, including 14 instead of 11 cysteines and a cluster of histidines in the C-terminal region, appear to be found exclusively in vertebrates.     

文昌鱼肝盲囊与脊椎动物肝脏起源

早期形态和胚胎学研究结果表明, 文昌鱼哈氏窝、内柱和肝盲囊分别是脊椎动物脑垂体、甲状腺和肝脏的同源器官。文章总结了近年来有关文昌鱼肝盲囊与脊椎动物肝脏之间关系的研究成果, 揭示文昌鱼肝盲囊和脊椎动物肝脏具有同源性, 并证明文昌鱼中存在类似脊椎动物的GH/IGF (Growth hormone/insulin-like growth factor)和TH/THR (Thyroid hormone/thyroid hormone receptor)信号通路, 为脊椎动物肝脏起源于文昌鱼肝盲囊样结构提供了分子水平证据。