High molecular weight DNA purification instrument for metagenomics

用于宏基因组学的高分子量DNA纯化仪

• 批准号: 7778514
• 负责人: ANDRE MARZIALI
• 金额: $ 11.29万
• 依托单位: UNIVERSITY OF BRITISH COLUMBIA
• 依托单位国家: 美国
• 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/7778514
• 关键词: Address; Antibiotics; Biochemical Reaction; Biological Markers; Body Fluids; Buffers; Centrifugation; Cloning; 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; Procedures; Process; Production; RNA Degradation; Reagent; Recovery; Research Personnel; Resources; Sampling; Site; Soil; Solutions; Speed; Staging; System; Techniques; Technology; Temperature; Testing; Vacuum; Work; bacterial lysate; base; commercialization; cost; design; electric field; inhibitor/antagonist; instrument; instrumentation; interest; macromolecule; microorganism; next generation; novel; process optimization; programs; prototype; reconstruction; simulation; soil sampling; tool

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.

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，因此 研究人员无法研究某些环境中 99% 以上的微生物 - 这些微生物 有时具有独特的能力，例如合成可用作新药或 抗生素。宏基因组学，即不可培养微生物的基因组重建，是一种强大的新方法 获取这些生物体未开发资源的工具。在一种方法中，DMA 的大片段 从含有不可培养微生物的样品中提取并克隆到大肠杆菌等宿主中 产生宏基因组库。然后筛选文库中表达的化合物的效用。 尽管用这种方法发现了新的抗生素和酶，但仍无法成功生产 由原始微生物合成的化合物，例如抗生素，提出了艰巨的挑战 足够长的克隆片段可以容纳基因簇编码的路径，这些基因簇通常超过 100kb 长度。现有的纯化技术倾向于将基因组 DMA 剪切为 50kb 或更小的片段。这 发现一种从土壤等困难样品中提取和纯化高分子量 DMA 的方法将为 宏基因组学的突破可能有助于发现许多未来的药物和抗生素。 我们最近展示了一种用于 DMA 浓度的非线性电泳方法，即 能够进行 DMA 操作和浓缩，无需机械处理（例如离心或 移液）。从细菌裂解液中提取 DNA，浓缩倍数为 4,000 倍 证明了。该方法是下一代 DNA 提取方法的绝佳候选方法 土壤或体液等复杂样品的方法。我们建议开发一种仪器来携带 提出了这种浓缩方法，验证了其在土壤中提取高分子量 DNA 的性能 用于宏基因组学研究的样本，旨在发现新药和抗生素。对此的许可 该技术将致力于解决其他引人注目的应用，包括从体内提取 DNA 用于癌症生物标志物和病原体检测的液体。

项目成果

An instrument for automated purification of nucleic acids from contaminated forensic samples.

用于从受污染的法医样品中自动纯化核酸的仪器。

• DOI: 10.1016/j.jala.2007.10.008
• 发表时间: 2008
• 期刊: JALA (Charlottesville, Va.)
• 作者: Broemeling,DavidJ;Pel,Joel;Gunn,DylanC;Mai,Laura;Thompson,JasonD;Poon,Hiron;Marziali,Andre
• 通讯作者: Marziali,Andre