UIDA GENE-TRANSFER AND EXPRESSION IN MAIZE MICROSPORES USING THE BIOLISTIC METHOD

The ability to recover male gametophyte derived plants, which is necessary to get transformed haploid plants, was verified for a hybrid of maize. Using the isolated microspore culture technique, a 9 x 10(-5) plant regeneration frequency was obtained. Maize microspores were bombarded with tungsten particles using a PDS He/1000 apparatus. GUS expression in the microspores was maximum with 1.1 mu m diameter tungsten microprojectiles for 1100 and 1350 psi helium pressures at a 6 cm distance between the launch point and the target cells. Increasing the amount of DNA coated on the microparticles from 1.66 to 4 mu g DNA/mg of particles allowed a two-fold and four-fold increase of the GUS-expressing microspore frequency for 1100 and 1350 psi helium pressure bombardment, respectively. Optimal concentration of solidifying agent in the bombardment support culture medium was found to be 1%. Cell density ranging from 25000 microspores/bombardmenl to 100000 microspores/boombardment did not affect the frequency of GUS-expressing microspores. Using these optimal conditions, the maximum frequency of GUS-expressing microspores was found to be about 9 x 10(-4), while maintaining an embryo formation frequency about 5 x 10(-4).