[Agrobacterium tumefaciens-mediated transformation of the white-rot basidiomycete, phanerochaete chrysosporium].
Agrobacterium tumefaciens has been used to transfer its T-DNA to filamentous fungus Phanerochaete chrysosporium. A DNA fragment containing recombinant hygromycin B phosphotransferase gene (rhyg(r)) which is controlled by a promoter from P. chrysosporium and the lignin peroxidase gene terminator was cut out from pCH6, and inserted into a plant binary plasmid pCAMB1300, resulted in the recombinant plasmid pCH6-1300. The plasmid pCH6-1300 was then transformed into Agrobacterium tumefaciens strain A208, which was used to infect conidia or protoplasts of P. chrysosporium. Sixteen hygromycin B-resistant P. chrysosporium colonies were obtained. The transformation frequency is about 8 - 9 transformants per 106 conidia or protoplasts, which had no significant difference with the values obtained from conidia or protoplasts. One of the transformants named AT18, whose colony morphology was different from the wild-type strain. The results from both PCR and Southern blot analysis of transformants demonstrated that the rHyg(r) gene was integrated into the host genome with single copy. This approach may not require any instrument and digestion of fungal cell walls. A . tumefaciens-facilitated transformation should be very useful for further studies of the molecular genetics of P. chrysosporium.
Li
Wei
W
College of Life science, Sichuan University, Chengdu, China.
Zhang
Yi-zheng
YZ
D016960:Agrobacterium tumefaciens / Q000235:genetics*; D001487:Basidiomycota / Q000235:genetics*; D010544:Peroxidases / Q000235:genetics; D020075:Phanerochaete / Q000235:genetics*; D017853:Phosphotransferases (Alcohol Group Acceptor) / Q000235:genetics; D010957:Plasmids; D014170:Transformation, Genetic
D010544:Peroxidases; C042858:lignin peroxidase; D017853:Phosphotransferases (Alcohol Group Acceptor); C063242:hygromycin-B kinase