Construction of BAC Libraries of Metazoans Occupying Key Phylogenetic Positions

基因组DNA大插入文库是发育、进化和遗传学比较研究的宝贵资源，有助于解决基因组如何进化、基因组进化如何导致人体计划进化等生物学基本问题。拟议研究的目标是构建11个代表后生动物广度的BAC文库。重点放在对理解动物进化至关重要的系统发育地位的动物上。优先选择具有发展潜力的生物作为发育模式生物。因此，我们选择了两种后口动物：七鳃鳗，颅底，对了解脊椎动物进化过程中的基因和染色体复制模式至关重要；头索文昌鱼，与脊椎动物最接近的活体无脊椎动物近亲。根据分子系统发育，原口动物最近被划分为蜕皮虫和光生毛虫两个类群。然而，这些类群中每一个类群的分枝模式都没有得到很好的解决。由于蜕皮虫、果蝇和线虫的代表性已经被很好地研究，我们没有选择任何蜕皮虫。对于一般对基因、发育和进化很少关注的细毛类动物，已经选择了7种生物。其中包括2种环节动物，其中一种是多毛类扁形动物，在环节动物中属于相对碱性的动物，另一种是水蚤，是一种高度进化的模式生物。扁形动物正在迅速成为一种模式生物，而水蚤是相当衍生的，多年来它一直是发育遗传学研究的对象。包括两种软体动物--一个双壳类(Spisula Solidissima)和一个脚类(Ilyansoliobeta)。其他的线虫包括1种线虫(Cerebratulus Lacteus)、1种管形吸虫(Temiste Langformis)和1种扁虫(NeoChildea Fusca)。关于无脊椎动物的系统发育位置存在争议。在形态的基础上，它们被认为是基本的双毛目。然而，大多数，但不是全部的，分子系统发育分析都将触角放在原口内。为了解决双毛目的起源，我们选择了一个鞭毛虫(Mnformopsis Leidyi)。最后，为了解决后生动物的起源问题，选择了一种海绵--扩散海绵，它是最基本的后生动物的代表。BAC文库将由P.de Jong(奥克兰儿童医院研究所)构建和分发，他在BAC文库的建设方面有着长期的记录，包括为人类基因组计划制作BAC文库。对于对整个基因组测序感兴趣的生物，建议覆盖15倍。对于其他人，建议覆盖10倍。

Large-insert libraries of genomic DNA are a valuable resource for comparative studies indevelopment evolution and genetics for addressing such fundamental questions in biology ashow genomes have evolved and how evolution of genomes has resulted in evolution of bodyplans. The goal of the proposed research is to construct 11 BAC libraries representative of thebreadth of the Metazoa. The focus is on animals that occupy phylogenetic positions key to anunderstanding of animal evolution. Preference is given to organisms with demonstrated potentialas developmental model organisms. Therefore, 2 deuterostomes have been chosen: lamprey,basal in the craniates and pivotal for understanding the pattern of gene and chromosomalduplications during vertebrate evolution, and the cephalochordate amphioxus, the closest livinginvertebrate relative of the vertebrates. On the basis of molecular phylogenetics, the protostomeshave recently been divided into two groups, the Ecdysozoa and the Lophotrochozoa. However,branching patterns of the phyla within each of these groups are poorly resolved. Becauserepresentatives of the Ecdysozoa, Drosophila and Caenorhabditis have been well-studied, wehave not chosen any Ecdysozoan. For the Lophotrochozoa, which have, in general, received littleattention in regard to genes, development and evolution, seven organisms have been chosen.These include 2 annelids, one of which, the polychaete Platynereis dumerilii, is relatively basalwithin the annelids and the other of which, the leech Helobdella robusta, is more highly derived.Platynereis is rapidly becoming a lophotrochozoan model developmental organism, and whilethe leech is quite derived, it has been the object of developmental genetic studies for some years.Two molluscs are included-- one bivalve, (Spisula solidissima) and one gastropod (Ilyanassaobsoleta). The other Lophotrochozoans include 1 nemertean (Cerebratulus lacteus), 1 sipunculid(Temiste langiformis), and 1 acoel flatworm (Neochildea fusca). The phylogenetic position ofthe acoels is controversial. On the basis of morphology, they were considered basal Bilateria.However, most, but not all, molecular phylogenetic analyses place acoels within the protostomes.To address bilaterian origins, we have chosen a ctenophore (Mnemiopsis leidyi). Finally, toaddress the origin of the metazoa, a sponge, Callyspongia diffusa, a representative of the mostbasal Metazoa, was selected. BAC libraries will be constructed and distributed by P. de Jong(Children's Hospital Research Institute, Oakland), who has a long track-record in construction ofBAC libraries, including making the BAC library for the human genome project. For organismsfor which interest is high to sequence the entire genome, 15X coverage is proposed. For others,10X coverage is proposed.