High molecular weight DNA purification instrument for metagenomics

用于宏基因组学的高分子量DNA纯化仪

基本信息

批准号：
7367898
负责人：
ANDRE MARZIALI
金额：
$ 20.58万
依托单位：
UNIVERSITY OF BRITISH COLUMBIA
依托单位国家：
美国
项目类别：
财政年份：
2006
资助国家：
美国
起止时间：
2006-03-01 至 2009-08-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/7367898
关键词：
Address Antibiotics Bacterial DNA Biochemical Reaction Biological Markers Body Fluids Buffers Centrifugation Cloning Compatible Complex Coupled Cytolysis DNA DNA purification Development Electrodes Electrophoresis Environment Enzymes Escherichia coli Frequencies Future Gel Gene Cluster Generations Genomics Grant Injection of therapeutic agent Laboratories Left Length Libraries Licensing Malignant Neoplasms Mechanics Metagenomics Methods Molecular Weight Nature Nucleic Acids One-Step dentin bonding system Organism Pathogen detection Pathway interactions Performance Pharmaceutical Preparations Polymerase Chain Reaction Procedures Process Production RNA Degradation Range Reagent Recovery Research Personnel Resources Sampling Site Soil Solutions Speed Staging System Techniques Technology Temperature Testing Vacuum Work bacterial lysate base commercialization cost design desire electric field inhibitor/antagonist instrument instrumentation interest macromolecule microorganism next generation novel programs prototype reconstruction simulation size soil sampling tool

项目摘要

Less than 1% of all microorganisms observed in nature can be cultured in the laboratory, leaving researchers unable to study more than 99% of microorganisms in some environments - microorganisms that sometimes have unique abilities such as synthesis of compounds that could find use as new drugs or antibiotics. Metagenomics, the genomic reconstruction of unculturable microorganisms , is a powerful new tool for accessing the untapped resources of these organisms. In one approach, large fragments ofDMA from a sample containing unculturable microorganisms are extracted and cloned into a host such as E. coli to produce a metagenomics library. The library is then screened for utility of expressed compounds. Though new antibiotics and enzymes have been discovered with this method, successful production of compounds, such as antibiotics, synthesized by the original microorganism presents the difficult challenge of cloning fragments long enough to hold pathways encoded by gene clusters that are often over 100kb in length. Existing purification techniques tend to shear genomic DMAto fragments of 50kb or less. The discovery of a method to extract and purify high MW DMAfrom difficult samples such as soil will provide a breakthrough for metagenomics that may enable the discovery of many future drugs and antibiotics. We have recently demonstrated a non-linear electrophoretic method for DMA concentration that is capable of DMA manipulation and concentration without mechanical handling (such as centrifugation or pipetting). Extraction of DNA from bacterial lysate and 4,000 fold concentration factors have been demonstrated. This method is an excellent candidate for the next generation of methods for DNA extraction methods from complex samples such as soil or body fluids. We propose to develop an instrument to carry out this concentration method, validating its performance on extraction of high molecular weight DNA on soil samples for metagenomics studies aimed at discovery of new drugs and antibiotics. Licensing of this technology will be pursued to address other compelling applications including DNA extraction from body fluids for cancer biomarker and pathogen detection.

在自然界中观察到的所有微生物中，只有不到1％可以在实验室中培养研究人员无法在某些环境中研究超过99％的微生物 - 微生物 - 有时具有独特的能力，例如合成化合物，这些化合物可以找到新药或抗生素。宏基因组学是无法培养的微生物的基因组重建，是一种有力的新事物用于访问这些生物的未开发资源的工具。在一种方法中，大的碎片ofdma 从包含不可培养的微生物的样品中提取并将其克隆到宿主中，例如大肠杆菌生产宏基因组学库。然后将库筛选以获取表达化合物的实用性。尽管通过这种方法发现了新的抗生素和酶，但成功生产由原始微生物合成的化合物，例如抗生素，提出了困难的挑战克隆片段的时间足够长，可以容纳通常超过100kb的基因簇编码的途径长度。现有的纯化技术倾向于剪切50kb或更少的基因组DMATO片段。这发现一种提取和净化高MW DMAFROM的方法（例如土壤）的方法将提供宏基因组学的突破，可能会发现许多未来的药物和抗生素。我们最近证明了一种非线性电泳方法，用于DMA浓度能够在没有机械处理的情况下进行DMA操纵和浓度（例如离心或移液器）。从细菌裂解物中提取DNA和4,000倍浓度因子已经是证明。该方法是下一代DNA提取方法的绝佳候选者来自土壤或体液等复杂样品的方法。我们建议开发一种携带的工具删除这种浓度方法，验证其在提取土壤上高分子重量DNA的表现旨在发现新药和抗生素的宏基因组学研究样本。许可将采用技术来解决其他引人注目的应用程序，包括从身体中提取DNA 癌症生物标志物和病原体检测的流体。