Cloning，Expression and Identification of Apiscerana cerana Acetylcholinesterase Gene in Pichia pastoris

Abstract:

Organophosphate（OP） pollution is a major hidden danger to food safety，acetylcholinesterase can be used to detect the residual OP pesticide. Apiscerana cerana is a unique kind of Apoidea which has not been found in other countries except for China，and it has a high sensitivity to pesticides. On the other hand，acetylcholinesterase gene from Apiscerana cerana has not been cloned and applied in food safety detection. In this study，using the total RNA from Apiscerana cerana head tissue as a template，a 1.8kb AChE cDNA sequence was obtained by RACE method，then the target sequence was cloned into the yeast expression vector pPIC3.5K. AChE was expressed and located in Pichia pastoris GS115 after induced with 0.5% methanol. The SDS-PAGE and activity analysis showed that Apiscerana cerana AChE gene was successfully cloned and expressed in Pichia pastoris for the first time. A strain with higher activity AChE was selected to express AChE，and its AChE activity was 10 568 U/mg，which could be used to detect the residual OP pesticide.