Chames Patrick, Epinat Jean-Charles, Guillier Sophie, Patin Amélie, Lacroix Emmanuel, Pâques Frédéric
CELLECTIS S.A., 102 route de Noisy 93235, Romainville, France.
Nucleic Acids Res. 2005 Nov 23;33(20):e178. doi: 10.1093/nar/gni175.
Homing endonucleases, endonucleases capable of recognizing long DNA sequences, have been shown to be a tool of choice for precise and efficient genome engineering. Consequently, the possibility to engineer novel endonucleases with tailored specificities is under strong investigation. In this report, we present a simple and efficient method to select meganucleases from libraries of variants, based on their cleavage properties. The method has the advantage of directly selecting for the ability to induce double-strand break induced homologous recombination in a eukaryotic environment. Model selections demonstrated high levels of enrichments. Moreover, this method compared favorably with phage display for enrichment of active mutants from a mutant library. This approach makes possible the exploration of large sequence spaces and thereby represents a valuable tool for genome engineering.
归巢内切酶，即能够识别长DNA序列的内切酶，已被证明是精确高效基因组工程的首选工具。因此，设计具有定制特异性的新型内切酶的可能性正在深入研究中。在本报告中，我们提出了一种基于切割特性从变体文库中选择巨型核酸酶的简单有效方法。该方法的优点是直接选择在真核环境中诱导双链断裂诱导同源重组的能力。模型选择显示出高水平的富集。此外，该方法与噬菌体展示相比，在从突变体文库中富集活性突变体方面表现良好。这种方法使得探索大的序列空间成为可能，因此是基因组工程的一种有价值的工具。
