OPTIMIZATION OF DNA TRANSFER AND TRANSIENT BETA-GLUCURONIDASE EXPRESSION IN ELECTROPORATED MAIZE (ZEA-MAYS L) MICROSPORES

The ability to deliver free DNA into microspores of a highly androgenic hybrid of maize was assessed by electroporation, using a square wave pulse discharge apparatus. The electroporation medium was chosen according to its ability to maintain a high level of regeneration. Nuclease activities were analyzed and were inhibited by the addition of 100 mM KNO3 and MgSO4 in the electroporation medium. Seven expression vectors with Uid A as the reporter gene under the control of cauliflower mosaic virus 35S, Lat 52-7, Zmg 13, Emu, Ubiq-1, Al, or Act1 promoters were tested in relation to the level of beta-glucuronidase expression in maize microspores. The highest level of expression was obtained when the Uid A gene was driven by the Act1 promoter. Therefore, this vector was further used to define optimal conditions leading to highest levels of beta-glucuronidase expression. The parameters determined in this study could provide an ideal starting point for the obtention of transgenic maize plants from electroporated microspores.