Agrobacterium-mediated transformation of the filamentous fungus Aspergillus awamori

被引：175
作者：
Michielse, Caroline B. [2 ]
Hooykaas, Paul J. J. [3 ]
van den Hondel, Cees A. M. J. J. [1 ]
Ram, Arthur F. J. [1 ]
机构：
[1] Leiden Univ, Inst Biol, Kluyver Ctr Genom Ind Fermentat, Dept Mol Microbiol, NL-2333 AL Leiden, Netherlands
[2] Univ Amsterdam, Swammerdam Inst Life Sci, Dept Plant Pathol, NL-1098 SM Amsterdam, Netherlands
[3] Leiden Univ, Dept Mol & Dev Genet, Inst Biol, NL-2333 AL Leiden, Netherlands
来源：
NATURE PROTOCOLS | 2008年 / 3卷 / 10期
关键词：
D O I：
10.1038/nprot.2008.154
中图分类号：
Q5 [生物化学];
学科分类号：
071010 ; 081704 ;
摘要：
Many transformation methods have been developed to introduce DNA into filamentous fungi. One of these methods is Agrobacterium-mediated transformation (AMT). Here, we describe an efficient protocol for AMT of Aspergillus awamori. This protocol has been used to determine the function of Agrobacterium virulence genes during AMT, to identify factors influencing transformation frequencies, to generate insertional mutants and to generate A. awamori gene knockout transformants. This protocol in not only applicable to A. awamori, but can be used as a more general guideline for AMT of other filamentous fungi. Conidiospores are incubated with induced Agrobacterium, and, after a cocultivation and selection period, hygromycin-resistant transformants are obtained with a frequency of 200-250 transformants per 1 x 10(6) conidiospores. Using this protocol, transformants can be obtained within 10-12 d.
引用
收藏
页码：1671 / 1678
页数：8
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