Gratz Andreas, Jose Joachim
Institute of Pharmaceutical and Medicinal Chemistry, Heinrich-Heine-University, Düsseldorf, Germany.
Methods Mol Biol. 2011;729:153-66. doi: 10.1007/978-1-61779-065-2_10.
Directed evolution is an often used approach toward new proteins with tailor-made properties. It consists of random variation of the coding sequence of a protein followed by an appropriate selection procedure or a suitable type of property read out. In many, if not all cases, it is of significant advantage to constrain the randomly mutagenized DNA sequence to that encoding a particular part of the protein or a distinct domain, and not to mutate the entire gene of the target protein. For this purpose, a three-step, polymerase-based method was developed, which is independent of two flanking restriction sites adjacent to the nucleotide sequence supposed to be mutagenized, and named protein library generation by overlap extension (PDLGO).
定向进化是一种常用于获得具有定制特性的新蛋白质的方法。它包括对蛋白质编码序列进行随机变异，然后进行适当的选择程序或读出合适类型的特性。在许多（如果不是所有）情况下，将随机诱变的DNA序列限制在编码蛋白质特定部分或不同结构域的序列上，而不是诱变目标蛋白质的整个基因，具有显著优势。为此，开发了一种基于聚合酶的三步法，该方法独立于与假定要诱变的核苷酸序列相邻的两个侧翼限制性位点，并将其命名为通过重叠延伸生成蛋白质文库（PDLGO）。
