IMPROVED SEPARATION MEDIA FOR ELECTROPHORESIS

改进的电泳分离介质

• 批准号: 6181625
• 负责人: Benjamin Chu
• 金额: $ 18.76万
• 依托单位: STATE UNIVERSITY NEW YORK STONY BROOK
• 依托单位国家: 美国
• 财政年份: 1995
• 资助国家: 美国
• 起止时间: 1995-09-30 至 2001-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6181625
• 关键词: DNA; analytical method; atomic force microscopy; birefringences; capillary electrophoresis; copolymer; crystallization; light scattering; low angle X ray diffraction analysis; method development; micelles; molecular assembly /self assembly; nonblood rheology; nuclear magnetic resonance spectroscopy; nucleic acid sequence; physical model; polymerization; reagent /indicator; solutions; viscosity

DESCRIPTION (Adapted from the Investigator's Abstract): The specific aim of the renewal proposal deals with further development and optimization of new and better separation media which can overcome many of the problems related to the present-day polyacrylamide solutions or gels: polymer solutions are often too viscous to be filled in capillary tubings; the polymerization in situ approach is not so reproducible; shelf-life for polymer gels is limited. The key scheme is to take advantage of the self-assembly behavior of copolymer colloids. The investigators have achieved the initial goals of designing a polymer network based on the self-assembly behavior of a EPE-type block copolymer (E and P denoted polyoxyethylene and polyoxypropylene, respectively) with the following properties: (1) at 10-15 degrees C, the polymer solution is a low viscosity fluid (less than 30 centipoise) in 1xTBE buffer, for ease of filling and removing the separation medium in narrow-bore capillary tubings; (2) at room temperatures (greater than 20 degrees C), the polymer solution becomes gel-like with good sieving ability (tested down to 14 bases with one base resolution for single stranded (ss) DNA fragments and up to 1 kilobase pairs for ds DNA fragments with one base pair resolutions); and (3) the polymer itself has acted successfully as a dynamic coating material. By correlating the capillary electrophoresis results with structural studies using a variety of physical techniques, such as laser light scattering, small angle X-ray scattering, small angle neutron scattering, NMR, rheometry, viscosimetry and atomic force microscopy, an optimization of the self-assembled polymer network for DNA capillary electrophoresis can be achieved. In addition to the EPE, PEP, EBE, and BEB type triblocks (with B being polyoxybutylene and more hydrophobic than P), it is proposed to use crystallization and coil-collapse as the physical phenomena capable of temperature-dependent reversible cross-linking formation. The crystallization approach is especially attractive, since the supramolecular structure can be more resilient and is not in dynamic equilibrium with single polymer chains as in the self-assembly process.

描述（改编自研究者的摘要）：具体目标 更新提案涉及新产品的进一步开发和优化 更好的分离介质可以克服许多相关问题 到现在的聚丙烯酰胺溶液或凝胶：聚合物溶液是 通常太粘稠而无法填充到毛细管中；聚合反应在 现场方法的可重复性较差；聚合物凝胶的保质期为 有限的。 关键方案是利用自组装行为 的共聚物胶体。 研究人员已经实现了设计聚合物的初步目标 基于 EPE 型嵌段共聚物（E P 分别表示聚氧乙烯和聚氧丙烯） 具有以下性能：（1）在10-15℃时，聚合物溶液具有低 1xTBE 缓冲液中的粘度液体（小于 30 厘泊），以便于 填充和去除窄孔毛细管中的分离介质； (2)在室温(大于20℃)下，聚合物溶液 变成凝胶状，具有良好的筛分能力（测试到 14 个碱基，其中一个 单链 (ss) DNA 片段的碱基分辨率高达 1 kilobase 具有一个碱基对分辨率的双链 DNA 片段对）；和（3） 聚合物本身已成功地用作动态涂层材料。 通过将毛细管电泳结果与结构研究相关联 使用各种物理技术，例如激光散射， 小角X射线散射、小角中子散射、NMR、 流变测定法、粘度测定法和原子力显微镜，优化 用于 DNA 毛细管电泳的自组装聚合物网络可以 实现了。 除了 EPE、PEP、EBE 和 BEB 型三嵌段（B 是聚氧化丁烯，比 P 更疏水），建议使用 结晶和线圈塌陷作为物理现象能够 温度依赖性可逆交联形成。 这 结晶方法特别有吸引力，因为超分子 结构可以更具弹性并且不处于动态平衡 自组装过程中的单聚合物链。