Agrobacterium mediated transformation of moulds, in particular those belonging to genus aspergillus

本发明涉及农杆菌介导的霉菌包括真菌子囊菌、担子菌、半知菌、鞭毛菌和接合菌亚部的霉菌种的转化。实施例证明了泡盛曲霉(原生质体和分生孢子)、构巢曲霉、黑曲霉、盘长孢状刺盘孢、豌豆腐皮镰孢、粗糙脉孢菌、雷西木霉、糙皮侧耳和二孢蘑菇(均为分生孢子)、以及禾本科镰孢(分生孢子和重新水化的ATCC冻干物)的转化。尤其在泡盛曲霉中，转化频率远高于传统的霉菌转化技术。还进一步发现，不仅一个可表达的基因能被导入这些霉菌，甚至这种基因的多个拷贝也可以，而且，如同用泡盛曲霉所举的例子，这种基因可被定向至诸如染色体的pyrG位点中。这些多拷贝可以是一种编码一种所需的、同源或异源的蛋白的基因。 The invention relates to Agrobacterium mediated transformation of moulds comprising species of the fungal sub-divisions Ascomycotina, Basidiomycotina, Deuteromycotina, Mastigomycotina, and Zygomycotina.Examples demonstrate the transformation of Aspergillus awamori (both protoplasts and conidia), Aspergillus nidulans, Aspergillus niger, Colletotrichum gloeosporioides, Fusarium solani pisi, Neurospora crassa, Trichoderma reesei, Pleurotus ostreatus and Agaricus bisporus (all conidia), and Fusarium graminearum (both conidia and rehydrated freeze dried ATCC material).Especially for Aspergillus awamori the transformation frequency is much higher than with conventional mould transformation techniques.It has further been found that not only one expressable gene can be introduced into these moulds, but even multiple copies of such gene, which, moreover, can be targeted e.g. in the chromosomal pyrG locus, as exemplified for A. awamori. These multiple copies can be of a gene encoding a desired, homologous or heterologous, protein.