Method
Constructs were made containing selected genes flanked by regions homologous to the Ganoderma lucidum gpd promoter. Thus, the construct would be compatible with homologous recombination. The construct was made using a three part gibson assembly. The three parts were all made in separate BioBricks containing the left and right homology regions and the selected genes. The genes inserted into the homology flanking regions include prtpc promoter, gpd promoter, T7 promoter, and the resistance genes hph and nptII. The constructs were then inserted into the respective fungus strain using protoplasting and PEG transformation methods [1].
References
1. Gouka RJ, Gerk C, Hooykaas PJJ, Bundock P, Musters W, Verrips CT, de Groot MJA (1999). Transformation of Aspergillus awamori by Agrobacterium tumefaciens-mediated homologous recombination. Nature Biotechnology, 17, 598-601.