Optimisation of transformation conditions and production of transgenic sorghum (Sorghum bicolor) via microparticle bombardment

Conditions for microparticle bombardment were optimised for four explant types of sorghum Sorghum bicolor (L.) Moench based on transient expression of the uidA reporter gene. The tested physical parameters included acceleration pressure, target distance, gap width and macroprojectile travel distance. The sorghum explants studied were immature and mature embryos, shoot tips and embryogenic calli. In addition, the activity of four heterologous promoters was determined both by GUS histochemical staining and enzymatic activity assay in immature embryos and shoot tips. The strength of these promoters could be placed in the following order: ubi1>act1D>adh1>CaMV 35S. The optimised bombardment conditions were applied for selecting phosphinothricin- or geneticin-resistant in vitro cultures in order to generate transgenic plants. Production of transgenic plants via phosphinothricin-selection was not successful due to the release of phenolic substances from the herbicide-resistant cultures during the regeneration process. After selection on geneticin, however, fertile transgenic sorghum plants were regenerated from immature embryos as well as from shoot tips. Stable integration and Mendelian inheritance of the neo selectable marker gene was demonstrated in all transgenic plants.