REU: Ribosomal RNA Sequences in Marine Yeasts: A Model for Identification and Quantification of Marine Eukaryotes

区分海洋微生物种类(细菌、原生动物、真菌和微藻)的能力是理解初级生产和有机循环过程的一个主要问题。分子生物学的发展为从基因上描述和量化海洋微生物提供了一系列工具。将这些技术应用于生物海洋学问题的第一步是开发物种特有的遗传标记。核糖体RNA(RRNA)的5S亚基、小亚基(SSU)和大亚基(LSU)已被广泛研究。对进化缓慢的rRNA保守区的研究对发展进化的系统发育线起到了重要作用。除了保守区外，LSU中还有一些进化迅速的可变区。在费尔博士的实验室和其他地方进行的初步研究表明，可变区的核苷酸序列足够保守，可以在物种和属的水平上区分生物。这项研究将利用海洋酵母作为其他海洋真核生物的模型，扩大对LSU rRNA可变区的初步研究。酵母是唯一合适的，因为它们可以在培养中生长，它们的核苷酸序列可以与已知的交配遗传标记进行比较。核苷酸测序将使用桑格和莱恩的既定技术。识别物种特有的核苷酸序列，结合聚合酶链式反应和使用分子探针和杂交等技术，为扩增核酸以及从单细胞材料中鉴定和量化海洋微生物提供了强有力的工具。

The ability to distinguish species of marine microorganisms (bacteria protozoans, fungi and microalgae) is a major problem in understanding the processes of primary production and organic recycling. Developments in molecular biology are providing an array of tools to genetically characterize and quantify marine microbes. The first step in applying these techniques to problems in biological oceanography is to develop species- specific genetic markers. There has been extensive study on the 5S, small (SSU) and large (LSU) subunits of ribosomal RNA (rRNA). Studies of conserved regions of rRNA, which evolve slowly, have been instrumental in developing phylogenetic lines of evolution. In addition to conserved regions, there are variable regions in LSU that evolve rapidly. Initial investigations, in Dr. Fell's laboratory and elsewhere, suggest that nucleotide sequences in the variable regions are sufficiently conserved to distinguish organisms at the level of species and genus. This research will amplify preliminary studies of the variable regions of LSU rRNA using marine yeasts as a model for other marine eukaryotes. The yeasts are uniquely appropriate as they can be grown in culture and their nucleotide sequences can be compared with known mating genetic markers. Nucleotide sequencing will use the established techniques of Sanger and Lane. Recognition of nucleotide sequences that are species specific, in combination with techniques such as the polymerase chain reaction and the use of molecular probes and hybridizations, provide powerful tools for the amplification of nucleic acids and the identification and quantification of marine microorganisms from single cell material.