MOLECULAR PHYLOGENY OF EUKARYOTES

描述：提供了以前的工作，主要来自PIS实验室 真核生物进化史的概貌。虽然大多数人 生物学家把重点放在了真核生物(多细胞)上 真核生物)，是真核生物谱系进化史的主要部分 栖息在树下的大量未被研究的群体中。在……里面 特别是，已知的真核生物最早的分支谱系是 无线粒体，对基因组结构知之甚少 在这些血统中。 为了解决其中一些问题，公安部提出了三个具体目标： (1)从碱基中获得大量的编码序列 基于A-T的编码序列浓缩策略在真核生物中的应用 组合物，(2)使用16S核糖体序列进一步分解 更深的分支真核生物的结构，以及(3)使用大的 决定树冠分枝顺序的核糖体序列 辐射。 来自分支最低的真核生物的序列一直是非常困难的 通过标准的基于同调的方法获得给定的极值 这些血统的分化。私家侦探的实验室注意到 绿微孢子虫的基因组很小，A-T很高 含量，因此需要至少2G/C碱基对的限制性内切酶 预计将优先削减编码序列。序列 以这种方式隔离的随机克隆显示，每100个这样的克隆中有40个 与后来分化的真核生物中的基因具有非常显著的相似性。 PI建议使用此方法对200-1000个随机克隆进行排序 来自S·路菲的。初步数据显示，至少有400个克隆可以 在出现重复命中之前进行筛选。这些检测到的基因将 作为确定完整序列的同源探针，具有 包括200-500个碱基的侧翼序列。 同样的编码序列丰富策略将应用于两个 另一种分枝较深的线虫，即Intephalitoson intesinalis 和蓝氏贾第鞭毛虫。PI还建议获得小核糖体 来自低等真核生物关键谱系的序列，并使用大的 核糖体序列，以解决冠状真核生物中的辐射。

DESCRIPTION: Previous work, largely from the PIs laboratory, is provided a broad picture of the evolutionary history of eukarytoes. While most biologists focus on the "crown" eukaryotes (the multicellular eukaryotes), the bulk of evolutionary history of the eukarytoic lineages resides in a vast number of understudied groups lower in the tree. In particular, the earliest known branching lineages of the eukarytoes are amitochondriate, and very little is known about the genomic structures of these lineages. To resolve some of these issues, the PI proposes three specific aims: (1) to obtain a large number of coding sequences from the basal eukaryotes using a coding-sequence enrichment strategy based on A-T composition, (2) to use 16S ribosomal sequences to further resolve the structure of the deeper branching eukaryotes, and (3) to use large ribosomal sequences to resolve the branching order of the crown radiation. Sequences from the lowest branching eukaryotes have been very difficult to obtain by standard homology-based approaches given the extreme divergence of these lineages. The PI's laboratory noted that the microsporita Spraguea lophii has a very small genome and very high A-T content, so that restriction enzymes requiring at least 2 G/C base pairs are expected to preferentially cut coding sequencing. Sequences of random clones isolated this way show that 40/100 such clones displayed highly significant similarities to genes in later-diverging eukaryotes. The PI proposes to use this approach to sequence 200-1000 random clones from S. lophii. Preliminary data suggests that at lest 400 clones can be screened before duplicate hits appear. Such detected genes will serve as homologous probes for determination of complete sequences, with 200-500 bp of flanking sequences included. This same coding sequence enrichment strategy will be applied with two other, deeply branching amitochondriates, Encephalitozoon intesinalis and Giardia lamblia. The PI also proposes to obtain small ribosomal sequences from key lineages in the lower eukaryotes and to use large ribosomal sequences to resolve the radiation in the crown eukaryotes.