A Method for Rapid Determination of Monomer Sequences in Linear Polymeric Molecules such as Nucleic Acids

一种快速测定核酸等线性聚合物分子中单体序列的方法

• 批准号: 9421831
• 负责人: Daniel Branton
• 金额: $ 5万
• 依托单位: Harvard University
• 依托单位国家: 美国
• 项目类别: Standard Grant
• 财政年份: 1994
• 资助国家: 美国
• 起止时间: 1994-09-15 至 1995-08-31
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=9421831&HistoricalAwards=false
• 关键词: Method; Rapid; Determination; Monomer; Sequences

9421831 Branton We propose a new method for rapidly sequencing polymers such as DNA or RNA. The method involves measurements of ionic current modulation as the nucleotides of a linear nucleic acid molecule pass through a channel in an artificial membrane. Our immediate goal is to prove that we can draw single stranded DNA or RNA through a continuously open channel in a lipid bilayer and to demonstrate that, during polymer passage through the channel, ionic currents are reduced in a manner that reflects the properties of the polymer (length, nucleotide composition, etc.). Our long term goals are to demonstrate the feasibility of using a patch-clamp- like approach for direct electro-sensing of monomer sequences in a linear polymer of DNA; and develop this as a routine procedure for sequencing. Because the approach proposed eliminates several of the currently required preparatory chemical steps in DNA sequencing and because it depends on inherently rapid, molecular events, successful sequencing with this method will be an extraordinarily important advance that should reduce the time and cost of nucleic acid sequencing by several orders of magnitude. %%% A new method for the rapid sequencing of nucleic acid polymers like DNA and RNA will be developed. The method to be used involves measurements of changes in ionic currents as the individual building blocks of a nucleic acid pass through a channel in an artificial lipid membrane. The immediate goal of this research project is to show that single stranded DNA or RNA can be drawn through a continuously open channel in the membrane (lipid bilayer) and to demonstrate that during passage of the polymer changes in ionic currents are reduced in a manner that reflects the properties of the nucleic acid (i.e., length, composition of the individual nucleotides which make up the nucleic acid). The long term goals are to demonstrate the feasibility of using this patch- clamp-like approach for direct electro-sensing of ind ividual nucleotides (the building blocks of nucleic acids) in the linear sequence of DNA; and to develop this as a routine procedure for DNA sequencing. Because the proposed approach eliminates several of the currently required preparatory chemical steps in DNA sequencing and because it depends on inherently rapid molecular events, successful sequencing with this method will be an important advance that should reduce the time and cost of nucleic acid sequencing by several orders of magnitude.

我们提出了一种快速测序聚合物如DNA或RNA的新方法。 该方法涉及当线性核酸分子的核苷酸通过人工膜中的通道时测量离子电流调制。 我们的近期目标是证明我们可以通过脂质双层中的连续开放通道绘制单链DNA或RNA，并证明在聚合物通过通道期间，离子电流以反映聚合物性质（长度，核苷酸组成等）的方式减少。 我们的长期目标是证明使用膜片钳样方法直接电感测DNA线性聚合物中的单体序列的可行性;并将其开发为测序的常规程序。 由于所提出的方法消除了DNA测序中目前所需的几个预备化学步骤，并且由于它依赖于固有的快速分子事件，因此使用该方法的成功测序将是一个非常重要的进步，其将减少核酸测序的时间和成本几个数量级。 将开发一种新的核酸聚合物（如DNA和RNA）快速测序方法。 所使用的方法涉及测量离子电流的变化，因为核酸的各个构件通过人工脂质膜中的通道。 该研究项目的直接目标是表明单链DNA或RNA可以通过膜（脂质双层）中的连续开放通道被吸引，并证明在聚合物通过期间，离子电流的变化以反映核酸性质的方式减少（即，组成核酸的单个核苷酸的长度、组成）。 长期目标是证明使用这种膜片钳样方法用于直接电感测DNA线性序列中的单个核苷酸（核酸的结构单元）的可行性;并将其开发为DNA测序的常规程序。 由于所提出的方法消除了DNA测序中目前所需的几个预备化学步骤，并且由于它依赖于固有的快速分子事件，因此使用该方法的成功测序将是一个重要的进步，其将减少核酸测序的时间和成本几个数量级。