Comparative DNA sequence analysis of the yeast genome

我们建议测定四种酵母菌的基因组序列(目标1)，以确定它们的DNA序列中保守的、因此可能的功能特征。与其专注于蛋白质和蛋白质编码DNA(这类研究的共同重点)，我们相反地寻求在非编码序列(目标2)中发现隐藏的功能特征，例如调节基因表达、染色体维护和编码非蛋白质编码RNA的序列。因为这些种类的序列进化很快，所以需要比较相对接近的物种的序列。我们得出的结论是，将酿酒酵母的DNA序列与至少3个其他酵母物种的DNA序列进行比对(即4向比对)将是观察到有意义的保守序列块所必需的。对4个不同物种的基因组进行全基因组鸟枪测序(2-3倍覆盖)，然后有限地填补4向比对中的空白，似乎是获得必要数据的最有效和最经济的方式。我们将通过测试一些保守序列(目标3)的功能，通过设计实验来识别转录调控序列及其结合的蛋白质，以及非蛋白质编码RNA，从而评估我们识别功能序列元件的能力。酵母菌是这些研究的优秀生物，因为1)它们相对较小的基因组使该项目具有成本效益，2)关于酵母的丰富信息可用于评估保守序列的重要性，3)可以设计深入的实验来测试保守序列的生物功能，以及4)这些数据将使研究这一重要模式生物的科学家的大社区受益。我们希望为比较DNA序列分析作为一种注释基因组的方法的发展做出贡献，并学习如何有效地使用它。

We propose to determine the genome sequences of four Saccharomyces species (Aim 1), for the purpose of identifying conserved, and therefore likely functional features in their DNA sequences. Rather than focusing on proteins and protein-coding DNA (a common emphasis of such studies), we seek instead to find hidden functional features in non-coding sequence (Aim 2), such as sequences regulating gene expression, chromosome maintenance, and encoding non-protein coding RNAs. Because these kinds of sequences evolve rapidly, sequences of relatively closely related species need to be compared. We have come to the conclusion that alignment of S. cerevisiae DNA sequence to that of at least 3 other Saccharomyces species (i.e., 4-way alignments) will be necessary to observe conserved blocks of sequence that are significant. Whole genome shotgun sequencing (2-3-fold coverage) of the genomes of 4 different species, followed by limited filling of gaps in the 4-way alignments, seems like the most efficient and economical way to acquire the necessary data. We will assess our ability to recognize functional sequence elements based on their conservation by testing the function of some of the conserved sequences (Aim 3) with experiments designed to identify transcriptional regulatory sequences and the proteins that bind to them, as well as non-protein coding RNAs. Yeasts are excellent organisms for these studies because 1) their relatively small genomes makes the project cost effective, 2) a wealth of information on yeast is available for assessing the significance of conserved sequences, 3) incisive experiments can be designed to test the biological function of conserved sequences, and 4) the data will benefit the large community of scientists studying this important model organism. We hope to contribute to the development of comparative DNA sequence analysis as an approach to annotating genomes, and learn how to use it effectively.