Yu Xinliang, Yu Yixiong
College of Chemistry and Chemical Engineering, Hunan Institute of Engineering, Donghu Road 18#, Xiangtan, Hunan, 411104, China,
Appl Biochem Biotechnol. 2014 Aug;173(8):2019-27. doi: 10.1007/s12010-014-0989-9. Epub 2014 May 27.
Non-Systematic Evolution of Ligands by EXponential enrichment (SELEX)selection of aptamers, a novel technology for aptamer selection from libraries of random DNA (or RNA) sequences, involves repetitive steps of partitioning without polymerase chain reaction (PCR) amplification between them. This selection is based on non-equilibrium capillary electrophoresis of equilibrium mixtures (NECEEM) and has exceptionally high efficiency. In this paper, a mathematical analysis was carried out to predict the levels of enrichment of non-SELEX selection under different conditions such as different protein concentrations and different efficiencies of partitioning. Investigated results suggest that the magnitude of the bulk affinity (k d) being 10(4) or 10(5) μM for the initial pool has no obvious effect on selective enrichment and that the first, second, and third rounds of non-SELEX selection have different optimum protein concentration values [T f] that give maximum enrichment levels when [T f] ranges from 0.0005 to 0.5 μM. The significance of analyzing selective enrichment of NECEEM-based non-SELEX with the efficiency of partitioning target-bound ligands from free ligands has been demonstrated.
指数富集的配体非系统进化(SELEX)筛选适体是一种从随机DNA(或RNA)序列文库中筛选适体的新技术,它涉及在不进行聚合酶链反应(PCR)扩增的情况下进行重复的分区步骤。这种筛选基于平衡混合物的非平衡毛细管电泳(NECEEM),具有极高的效率。本文进行了数学分析,以预测在不同条件下,如不同蛋白质浓度和不同分区效率下非SELEX筛选的富集水平。研究结果表明,初始文库的整体亲和力(kd)大小为10⁴或10⁵μM对选择性富集没有明显影响,并且非SELEX筛选的第一轮、第二轮和第三轮具有不同的最佳蛋白质浓度值[Tf],当[Tf]在0.0005至0.5μM范围内时可给出最大富集水平。分析基于NECEEM的非SELEX从游离配体中分离靶结合配体的效率对选择性富集的意义已得到证明。