SELEXION - SYSTEMATIC EVOLUTION OF LIGANDS BY EXPONENTIAL ENRICHMENT WITH INTEGRATED OPTIMIZATION BY NONLINEAR-ANALYSIS
SELEXION - SYSTEMATIC EVOLUTION OF LIGANDS BY EXPONENTIAL ENRICHMENT WITH INTEGRATED OPTIMIZATION BY NONLINEAR-ANALYSIS
复制标题
DOI：
10.1016/0022-2836(91)90509-5
复制
发表时间：
1991-12-05
影响因子：
5.6
通讯作者：
GOLD, L
中科院分区：
文献类型：
--
作者：
IRVINE, D;TUERK, C;GOLD, L
最近，有报道了分离具有特定生物活性的核酸分子的新技术。在每一种情况下，浓缩过程都包括重复几轮从复杂的核酸序列混合物中进行选择，然后对以所需方式发挥作用的配体序列进行聚合酶链式反应(PCR)扩增。实验条件的特殊变化会极大地改变这些过程的结果。在这项研究中，我们使用数学分析和计算机模拟来预测哪些变化具有最大的影响，并制定提高有效性的策略和指导方针。首先，我们进行重建测试，以证明基于平衡结合的数学描述足以解释仅几轮后在实验室获得的高水平浓缩。然后，我们展示了在广泛的条件下预期的浓缩；最后，我们确定了用于最大浓缩的最佳蛋白质和核酸浓度，同时还确保了即使是结合良好的稀有分子也有很高的可能性回收。增强有效性的策略和指南一般适用于DNA、RNA或多肽配体的系统浓缩过程，并已在交互式模拟程序中实施，用于使用任何感兴趣的目标进行综合非线性优化浓缩。
Recently, novel technologies for isolation of nucleic acid molecules with specific biological activities have been reported. In each case, the enrichment process involves repeated rounds of selection from complex mixtures of nucleic acid sequences, followed by polymerase chain reaction (PCR) amplification of ligand sequences that function in the desired manner. Particular variations in experimental conditions can dramatically alter the outcome of these processes. In this study, we use mathematical analysis and computer simulation to predict which variations have the greatest impact and to develop strategies and guidelines for enhanced effectiveness. First, we perform reconstruction tests to demonstrate that a mathematical description based on equilibrium binding is sufficient to explain the high levels of enrichment attained in the laboratory after just a few rounds. Then, we show the expected enrichment for an extensive range of conditions; and, finally, we determine the optimum protein and nucleic acid concentrations to use for maximum enrichment, while also ensuring a high likelihood of recovering even the rare molecule that binds well. The strategies and guidelines for enhanced effectiveness are generally applicable to processes for systematic enrichment of DNA, RNA or peptide ligands and have been implemented in an interactive simulation program for integrated non-linear optimization of enrichment using any target of interest.