Agrobacterium tumefaciens-transient genetic transformation of Habanero pepper (Capsicum chinense Jacq.) leaf explants

Most of the pepper species of the genus Capsicum have been recalcitrant to efficient Agrobacterium tumefaciens -mediated stable or transient, genetic transformation. In the present work, we optimized a protocol for transient transformation of the Habanero pepper ( Capsicum chinense Jacq.) through the standardization of several experimental factors. These included the age of the plants, the temperature, the length of co-cultivation, the application of a negative (vacuum) and/or a positive (infiltration) pressure, along with micro injection, the use of acetosyringone during the bacterial culturing, and modification of the pH during the GUS assay to eliminate the endogenous β-glucuronidase activity. The standardized protocol, which yielded nearly 55% fully transformed leaf explants, was used to successfully mobilize two empty binary vectors (pCAMBIA2301 and pCAMex), as well as the C. chinense cDNAs encoding the pathogenesis-related protein 10 and esterase, respectively.