Levine Howard A, Nilsen-Hamilton Marit
Department of Mathematics, Iowa State University, Ames, IA 50011, United States.
Comput Biol Chem. 2007 Feb;31(1):11-35. doi: 10.1016/j.compbiolchem.2006.10.002. Epub 2007 Jan 10.
Systematic evolution of ligands by exponential enrichment (SELEX) is a procedure by which a mixture of nucleic acids that vary in sequence can be separated into pure components with the goal of isolating those with specific biochemical activities. The basic idea is to combine the mixture with a specific target molecule and then separate the target-NA complex from the resulting reaction. The target-NA complex is then separated by mechanical means (for example by filtration), the NA is then eluted from the complex, amplified by polymerase chain reaction (PCR) and the process repeated. After several rounds, one should be left with a pool of [NA] that consists mostly of the species in the original pool that best binds to the target. In Irvine et al. [Irvine, D., Tuerk, C., Gold, L., 1991. SELEXION, systematic evolution of nucleic acids by exponential enrichment with integrated optimization by non-linear analysis. J. Mol. Biol. 222, 739-761] a mathematical analysis of this process was given. In this paper we revisit Irvine et al. [Ibid]. By rewriting the equations for the SELEX process, we considerably reduce the labor of computing the round to round distribution of nucleic acid fractions. We also establish necessary and sufficient conditions for the SELEX process to converge to a pool consisting solely of the best binding nucleic acid to a fixed target in a manner that maximizes the percentage of bound target. The assumption is that there is a single nucleic acid binding site on the target that permits occupation by not more than one nucleic acid. We analyze the case for which there is no background loss (no support losses and no free [NA] left on the support). We then examine the case in which such there are such losses. The significance of the analysis is that it suggests an experimental approach for the SELEX process as defined in Irvine et al. [Ibid] to converge to a pool consisting of a single best binding nucleic acid without recourse to any a priori information about the nature of the binding constants or the distribution of the individual nucleic acid fragments.
指数富集配体的系统进化(SELEX)是一种程序,通过该程序,可将序列不同的核酸混合物分离成纯组分,目的是分离出具有特定生化活性的核酸。其基本思路是将混合物与特定靶分子结合,然后从所得反应中分离出靶 - 核酸复合物。接着通过机械手段(例如过滤)分离靶 - 核酸复合物,然后从复合物中洗脱核酸,通过聚合酶链反应(PCR)进行扩增,并重复该过程。经过几轮后,应该会得到一组核酸,其中大部分是原始混合物中与靶标结合最佳的物种。在欧文等人的研究中[欧文,D.,图尔克,C.,戈尔德,L.,1991年。SELEXION,通过指数富集和非线性分析进行综合优化的核酸系统进化。《分子生物学杂志》222卷,739 - 761页]对这一过程进行了数学分析。在本文中,我们重新审视欧文等人的研究[出处同上]。通过重写SELEX过程的方程,我们大幅减少了计算核酸组分逐轮分布的工作量。我们还建立了SELEX过程收敛到仅由与固定靶标结合最佳的核酸组成的库的充要条件,且这种收敛方式能使结合靶标的百分比最大化。假设靶标上有一个单一的核酸结合位点,且该位点最多只能被一个核酸占据。我们分析了无背景损失(无载体损失且载体上无游离核酸)的情况。然后我们研究存在此类损失的情况。该分析的意义在于,它为欧文等人[出处同上]所定义的SELEX过程提出了一种实验方法,使其能够收敛到由单一最佳结合核酸组成的库,而无需借助任何关于结合常数性质或单个核酸片段分布的先验信息。