An Integrated lab-on-chip system for genome sequencing of single microbial cells

用于单个微生物细胞基因组测序的集成芯片实验室系统

基本信息

批准号：
7938069
负责人：
Yu-Hwa Lo
金额：
$ 57.14万
依托单位：
UNIVERSITY OF CALIFORNIA, SAN DIEGO
依托单位国家：
美国
项目类别：
财政年份：
2008
资助国家：
美国
起止时间：
2008-09-26 至 2013-07-31
项目状态：
已结题

项目摘要

DESCRIPTION (provided by applicant): There are approximately 100 trillion microorganisms inhabited in a human body. The intimate interactions between these microorganisms and the host have a profound impact of the physiology of the human body. The majority of these microorganisms have yet to be fully characterized, mainly due to the difficulties in growing them in laboratory conditions. Here we propose the development of an efficient and scalable method to obtain genome sequences from single microbial cells. This will eliminate the need to obtain pure laboratory culture, thus allow us to systematically characterize the genome structure of microbial communities that resides in different parts of the human body. The proposed project contains three major components. 1. Development of low-cost and disposable cell sorting devices that integrate microfluidics with micro- scale optical components. Such a lab-on-chip cell sorter will be able to identify and isolate single microbial cells from samples that contain hundreds to thousands of different species and often are contaminated with free DNAs from the host and other sources. 2. Development of a micro-well based polymerase cloning device for simultaneous amplification of genomes from hundreds to thousands of single microbial cells in parallel. Using such a device, we will prepare sufficient amount of DNAs from single cells for whole genome shotgun sequencing, a critical step to obtain genome sequences from single cells. 3. Development of an integrate pipeline for dissecting the genome composition of human microbiome at the single cell resolution. We will test this pipeline by isolating and sequencing genome from approximately 35 single microbial cells at different levels of relative abundance from the mouse distal intestine. The propose method will provide the research community a new tool to identify unknown microbial species, to study their metabolic functions and to better understand the host-microbe interactions under various physiological and disease conditions. PUBLIC HEALTH RELEVANCE: The distribution and activities of microorganisms in the human body have a profound impact on the health of the host. In this proposed project we will develop an efficient and scalable method to obtain genome sequences from single cells in complex microbial communities. Characterizing the genomic composition and structures of these microorganisms is a necessary step before their metabolic functions and the interactions with the host can be study in great detailed.

描述（由申请人提供）：人体中居住着大约100万亿微生物。这些微生物与宿主之间的紧密相互作用对人体生理学产生了深远的影响。这些微生物中的大多数尚未完全表征，这主要是由于在实验室条件下生长它们的困难。在这里，我们提出了一种有效且可扩展的方法的发展，以从单个微生物细胞中获得基因组序列。这将消除获得纯实验室文化的需求，从而使我们能够系统地表征存在于人体不同部位的微生物群落的基因组结构。拟议的项目包含三个主要组成部分。 1。开发低成本和一次性细胞分选设备，这些设备将微流体与微尺度的光学成分相结合。这种实验室芯片细胞分选仪将能够从包含数百至数千种不同物种的样品中识别和分离单个微生物细胞，并经常被宿主和其他来源的游离DNA污染。 2。开发基于微孔的聚合酶克隆装置，用于同时扩增基因组从数百个单个微生物细胞并联。使用这样的设备，我们将准备足够数量的单个细胞中的DNA进行整个基因组shot弹枪测序，这是从单个细胞中获得基因组序列的关键步骤。 3。在单细胞分辨率下开发集成管道以剖析人类微生物组的基因组组成。我们将通过与小鼠远端肠相的相对丰度不同的大约35个单微生物细胞中分离和测序基因组来测试该管道。提出的方法将为研究社区提供一种新的工具，以识别未知的微生物物种，研究其代谢功能，并更好地理解各种生理和疾病条件下的宿主微生物相互作用。公共卫生相关性：微生物在人体中的分布和活动对宿主的健康有深远的影响。在这个拟议的项目中，我们将开发一种有效且可扩展的方法，以获取复杂微生物群落中单个细胞的基因组序列。表征这些微生物的基因组组成和结构是其代谢功能之前的必要步骤，并且可以详细研究与宿主的相互作用。