Institute of Entomology, Biology Centre ASCR, and Faculty of Science, University of South Bohemia, 37005 Ceske Budejovice, Czech Republic.
Insect Biochem Mol Biol. 2013 Jan;43(1):17-23. doi: 10.1016/j.ibmb.2012.10.011. Epub 2012 Nov 8.
Engineered nucleases are proteins that are able to cleave DNA at specified sites in the genome. These proteins have recently been used for gene targeting in a number of organisms. We showed earlier that zinc finger nucleases (ZFNs) can be used for generating gene-specific mutations in Bombyx mori by an error-prone DNA repair process of non-homologous end joining (NHEJ). Here we test the utility of another type of chimeric nuclease based on bacterial TAL effector proteins in order to induce targeted mutations in silkworm DNA. We designed three TAL effector nucleases (TALENs) against the genomic locus BmBLOS2, previously targeted by ZFNs. All three TALENs were able to induce mutations in silkworm germline cells suggesting a higher success rate of this type of chimeric enzyme. The efficiency of two of the tested TALENs was slightly higher than of the successful ZFN used previously. Simple design, high frequency of candidate targeting sites and comparable efficiency of induction of NHEJ mutations make TALENs an important alternative to ZFNs.
工程化核酸酶是能够在基因组的特定位点切割 DNA 的蛋白质。这些蛋白质最近已被用于许多生物体中的基因靶向。我们之前表明，锌指核酸酶（ZFNs）可以通过易错的非同源末端连接（NHEJ）DNA 修复过程用于在 B. mori 中产生基因特异性突变。在这里，我们测试了另一种基于细菌 TAL 效应蛋白的嵌合核酸酶在诱导家蚕 DNA 靶向突变中的效用。我们针对先前由 ZFN 靶向的基因组座 BmBLOS2 设计了三个 TAL 效应核酸酶（TALENs）。所有三个 TALEN 都能够在家蚕生殖细胞中诱导突变，表明这种嵌合酶的成功率更高。两种测试的 TALEN 的效率略高于先前使用的成功 ZFN。简单的设计、高频率的候选靶向位点和 NHEJ 突变诱导的可比效率使 TALEN 成为 ZFN 的重要替代物。
