Hu Yang, Dai Qingqing, Liu Yangyang, Yang Zhe, Song Na, Gao Xiaoning, Voegele Ralf Thomas, Kang Zhensheng, Huang Lili
State Key Laboratory of Crop Stress Biology for Arid Areas, College of Plant Protection, Northwest A&F University, Yangling, 712100, People's Republic of China.
Curr Microbiol. 2014 Jun;68(6):769-76. doi: 10.1007/s00284-014-0541-8. Epub 2014 Feb 20.
Valsa mali var. mali (Vmm), which is the causative agent of Valsa canker of apple tree, causes heavy damage to apple production in eastern Asia. In this article, we report Agrobacterium tumefaciens-mediated transformation (ATMT) of Vmm and expression of gfp (green fluorescent protein) in this fungus. The transformation system was optimized to a transformation efficiency of approximately 150 transformants/10(6) conidia, and a library containing over 4,000 transformants was generated. The tested transformants were mitotically stable. One hundred percent hph (hygromycin B phosphotransferase) integration into Vmm was identified by PCR and five single-copy integration of T-DNA was detected in the eighteen transformants by Southern blot. To our knowledge, this is the first report of ATMT of Vmm. Furthermore, this library has been used to identify genes involved in the virulence of the pathogen, and the transformation system may also be useful to the transformation of other species of the genus Valsa.
苹果黑腐皮壳菌苹果变种（Valsa mali var. mali，Vmm）是苹果树腐烂病的病原菌，给东亚地区的苹果生产造成了严重损失。在本文中，我们报道了根癌农杆菌介导的Vmm转化以及绿色荧光蛋白（gfp）在该真菌中的表达。转化系统优化后的转化效率约为150个转化子/10⁶分生孢子，并构建了一个包含4000多个转化子的文库。经检测，这些转化子在有丝分裂过程中是稳定的。通过PCR鉴定出hph（潮霉素B磷酸转移酶）100%整合到Vmm中，通过Southern杂交在18个转化子中检测到5个T-DNA单拷贝整合。据我们所知，这是关于Vmm根癌农杆菌介导转化的首次报道。此外，该文库已用于鉴定与病原菌毒力相关的基因，并且该转化系统可能对苹果黑腐皮壳菌属其他物种的转化也有用。
