Patterns of inheritance within the ciliate species Chilodonella uncinata - a poss

描述（由申请人提供）：本提案旨在阐明单一纤毛虫物种内的遗传模式，以解决三个特定的假设。这些假说集中在（1）体细胞（大核）基因组和（2）生殖细胞（微核）基因组内容的变化，以及（3）表观遗传学在性接合后形成体细胞基因组的可能作用。这些假设是基于两个分离株（一个来自美国，一个来自波兰）的一个单一形态种，Chilodonella uncinata的初步数据。虽然分离株含有相同的核糖体DNA，并共享一些相同的蛋白质编码序列，但它们也显示出复杂的遗传模式。基因的重复拷贝在接合后的大核中差异遗传，并且品系之间的杂交改变至少一个基因肌动蛋白的遗传模式。从这个项目中得到的数据将揭示纤毛Chilodonella uncinata内的遗传模式，以及可能与其他真核生物中的事件类似甚至同源的表观遗传过程。 为了解决这些假设，所提出的工作将结合联合收割机产生克隆文库，qPCR，变异特异性PCR和荧光显微镜。这些方法要么已经在PI实验室中使用，要么在荧光显微镜的情况下，目前正在开发中。 最后，拟议的工作将使PI继续参与学生（本科生和研究生）在她的研究计划。PI在从传统上代表性不足的群体中招募学生方面有着良好的记录，并将继续这些努力作为拟议工作的一部分。纤毛虫的拟议工作与公共卫生有关，因为由此产生的发现将深入了解真核细胞（有核细胞）中发育调控的基因组加工。在这项提议中要研究的基因组加工类型类似于人类免疫细胞和某些类型癌症中基因组的变化。对微生物真核生物（包括纤毛虫）的研究已经发现了许多与人类具有广泛相关性的基因组特征，包括染色体末端端粒的存在和自我剪接RNA的存在。

DESCRIPTION (provided by applicant): This proposal aims to elucidate patterns of inheritance within a single species of ciliates to address three specific hypotheses. These hypotheses focus on variation in the content of (1) somatic (macronuclear) genomes and (2) germline (micronuclear) genomes, and on (3) the possible role of epigenetics in shaping the generation of somatic genomes following sexual conjugation. These hypotheses are based on preliminary data from two isolates (one from the USA and one from Poland) of a single morphospecies, Chilodonella uncinata. Although the isolates contain identical ribosomal DNAs and share some identical protein coding sequences, they also show complex patterns of inheritance. Duplicated copies of genes are differentially inherited in macronuclei following conjugation and cross between lines alter patterns of inheritance of at least one gene, actin. The resulting data from this project will shed light on patterns of inheritance within the ciliate Chilodonella uncinata, and also on epigenetic processes that are likely analogous, and perhaps even homologous, to events in other eukaryotes. To address these hypotheses, the proposed work will combine generation of clone libraries, qPCR, variant-specific PCR, and fluorescent microscopy. These methods are either already in use in the PIs laboratory or, in the case of fluorescent microscopy, currently under development. Finally, the proposed work will enable the PI to continue to involve students (both undergraduate and graduate) in her research program. The PI has a demonstrated record in recruiting students from traditionally underrepresented groups, and will continue with these efforts as part of the proposed work. The proposed work on ciliates is relevant to public health in that resulting discoveries will yield insights into developmentally-regulated genome processing in eukaryotic cells (cells with nuclei). The type of genome processing to be studied in this proposal is analogous to changes in genomes in human immune cells and in some types of cancers. Studies of microbial eukaryotes, including ciliates, have already led to discovery of numerous genome features that have broad relevance to humans, including the presence of telomeres on the end of chromosomes and the existence of self-splicing RNA.