Highly efficient gene targeting in the Aspergillus niger kusA mutant
Highly efficient gene targeting in the Aspergillus niger kusA mutant
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DOI：
10.1016/j.jbiotec.2006.12.021
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发表时间：
2007-03-10
影响因子：
4.1
通讯作者：
Ram, Arthur F. J.
中科院分区：
文献类型：
--
作者：
Meyer, Vera;Arentshorst, Mark;Ram, Arthur F. J.
尼日尔中的基因靶向频率通常非常低，并且阻碍了这种生物技术上重要的真菌的有效功能基因组学。删除A.尼日尔kusA基因编码Ku70蛋白在其他真核生物中的直向同源物，当使用500 bp同源侧翼时，显著提高了同源整合效率，达到80%以上，而野生型背景中为7%。此外，在破坏必需基因的情况下，Delta kusA菌株的使用导致初级转化体中异核体形成的高频率（70%）。kusA的缺失对真菌的生长没有明显的影响，但使Delta kusA菌株对X射线照射的敏感性增加10倍，对紫外线照射的敏感性增加2至3倍。高效的基因打靶与A.尼日尔基因组序列允许系统的方法产生基因敲除，并将有助于提高A.尼日尔作为商业上感兴趣的蛋白质和代谢物的生产商。(c)2007 Elsevier B.V.保留所有权利。
Gene targeting frequencies in Aspergillus niger are often very low and hamper efficient functional genomics in this biotechnologically important fungus. Deletion of the A. niger kusA gene encoding the ortholog of the Ku70 protein in other eukaryotes, dramatically improved homologous integration efficiency and reached more than 80% compared to 7% in the wild-type background, when 500 bp homologous flanks were used. Furthermore, the use of the Delta kusA strain resulted in a high frequency of heterokaryon formation (70%) in primary transformants in the case disrupting an essential gene. Deletion of kusA had no obvious effect on the growth of the fungus, but renders the Delta kusA strain 10 times more sensitive to X-ray irradiation and two to three times more sensitive to UV exposure. The highly efficient gene targeting in combination with the A. niger genome sequence allows a systematic approach to generate gene knockouts and will help in improving the capacities of A. niger as producer of commercially interesting proteins and metabolites. (c) 2007 Elsevier B.V. All rights reserved.