Efficiency of Agrobacterium transformation of pinene synthase gene isolated from kaffir lime in Arabidopsis thaliana

Abstract

β-pinene, a monoterpene, was found as a major component of an essential oil extracted from fruit peel of kaffir lime (Citrus hystrix) known as Makrut. In this study, the pinene synthase gene (PNS) previously cloned from kaffir lime was constructed into a plant expression vector, pCAMBIA1305.1, and transformed into Arabidopsis thaliana. The kaffir lime PNS was replaced uidA (GUS) gene which was driven by CaMV35S promoter in the pCAMBIA1305.1 vector containing hygromycin resistance gene. After testing for appropriate hygromycin concentration, the PNS transgenic Arabidopsis mediated by Agrobacterium tumefaciens EHA105 were selected on 1/2MS containing 20 mg/l hygromycin for 10 days. The Arabidopsis seeds were collected from each transgenic lines of the 1^st–4^th generation (T0–T3) selected for hygromycin resistant. After selection, the percentage of seed survival of T0, T1, T2 and T3 transgenic Arabidopsis gradually increased at rate of 0.27, 45.17, 70.65 and 78.88, respectively. In addition, the kaffir lime PNS transgene was successfully integrated into the genome of all transgenic lines when detected by PCR. The obtained transgenic lines will be subjected to further study on the effects of this gene product to Arabidopsis essential oil components.