Circular polymerase extension cloning of cDNA glucoamylase Aspergillus awamori into integrative vector for filamentous fungi

This article describes successful insertion the cDNA glucoamylase from Aspergillus awamori into the integrative vector for filamentous fungi pH4Hyg using the CPEC (circular polymerase extension cloning) strategy. The prior step was the PCR amplification of the cDNA glaA gene cloned into a vector based on the plasmid pBluescript II SK(-) and the amplification of linear pH4Hyg vector with promotor and terminator of the glaA gene. The PCR products were purified and used for CPEC reaction. The CPEC product was formed after 20 cycles of the reaction. It is proposed to use the final plasmid pH4HygPTgla in engineering of filamentous fungi strains producing stable forms of glucoamylase with biotechnological interest.