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QPASS: Quantitative Parallel Aptamer Selection System

QPASS：定量平行适体选择系统

基本信息

批准号：
8520299
负责人：
LLOYD M SMITH
金额：
$ 99.98万
依托单位：
UNIVERSITY OF CALIFORNIA SANTA BARBARA
依托单位国家：
美国
项目类别：
财政年份：
2011
资助国家：
美国
起止时间：
2011-09-25 至 2015-01-31
项目状态：
已结题

项目摘要

Nucleic acid aptamers possess many useful features as affinity reagents, including facile chemical synthesis, reversible folding, thermal stability and low cost, making them a powerful alternative to antibodies and other protein-based reagents. However, over the past two decades, aptamers have suffered from the fact that 1) the conventional method of aptamer generation (SELEX) is lengthy, labor intensive and often does not yield aptamers with sufficient affinity (< 1 nM) and specificity; 2) there is no "standard protocol" that can be generally applied to most protein targets to generate aptamers; and 3) the characterization steps to measure the affinity and specificity of candidate aptamers are lengthy and resource-intensive, because each aptamer must be measured individually. We believe that these challenges arise from deficiencies in the conventional methodology of performing the selection, which has not changed significantly since its initial description 20 years ago. We also believe that these problems can be solved, by systematically taking fundamentally different approaches towards the three central stages of the process - selection, analysis and characterization of the aptamers. We propose here the development of such a system. We will combine three distinctly novel technologies -microfluidic selection, next-generation aptamer sequencing, and SPR Imaging - to develop the Quantitative Parallel Aptamer Selection System (QPASS) platform. The QPASS platform will generate specific aptamers with sub-nanomolar affinities (Kd) for a wide range of protein targets within 3 rounds of selection, identify a pool of the best candidates by next generation DNA sequencing and bioinformatic analysis, and home in on the optimal aptamer sequence by the parallel synthesis and measurement of the affinities of thousands of aptamer candidates. Individually, each component represents a significant technological advance. Combined this integrated approach offers an opportunity to revolutionize the process of aptamer generation.

核酸适体作为亲和试剂具有许多有用的特征，包括容易的化学合成、可逆折叠、热稳定性和低成本，使其成为抗体和其他基于蛋白质的试剂的有力替代品。然而，在过去的二十年中，适体受到以下事实的困扰：1）适体产生的常规方法（SELEX）是冗长的、劳动密集型的，并且通常不能产生具有足够亲和力（< 1 nM）和特异性的适体; 2）没有通常可以应用于大多数蛋白质靶以产生适体的“标准方案”;和3）测量候选适体的亲和力和特异性的表征步骤是冗长和资源密集的，因为必须单独测量每个适体。我们认为，这些挑战是由于进行甄选的传统方法存在缺陷造成的，而这种方法自20年前首次描述以来没有发生重大变化。我们还相信，这些问题可以通过系统地对该过程的三个中心阶段--适体的选择、分析和表征--采取根本不同的方法来解决。我们在此建议开发这样一个系统。我们将结合联合收割机三种独特的新技术-微流体选择，下一代适体测序和SPR成像-开发定量平行适体选择系统（QPASS）平台。QPASS平台将在3轮选择内生成对广泛蛋白质靶标具有亚纳摩尔亲和力（Kd）的特异性适体，通过下一代DNA测序和生物信息学分析鉴定最佳候选物的库，并通过平行合成和测量数千种适体候选物的亲和力来确定最佳适体序列。每个组件都代表着一项重大的技术进步。结合这种整合方法提供了一个机会，革命性的适体产生的过程。

项目成果

期刊论文数量（4）

专著数量（0）

科研奖励数量（0）

会议论文数量（0）

专利数量（0）

Fabrication of oligonucleotide and protein arrays on rigid and flexible substrates coated with reactive polymer multilayers.

DOI：
10.1021/am302285n
发表时间：
2013-01-23
期刊：
ACS APPLIED MATERIALS & INTERFACES
影响因子：
9.5
作者：
Broderick, Adam H.;Carter, Matthew C. D.;Lockett, Matthew R.;Smith, Lloyd M.;Lynn, David M.
通讯作者：
Lynn, David M.

In Vitro Selection of pH-Activated DNA Nanostructures.