Gene Transfer Into Intact Sugarcane Cells Using Microprojectile Bombardment

Abstract

A microprojectile accelerator has been constructed and used to bombard cultured sugarcane tissues with GUS reporter gene constructs. Design features useful to minimise target tissue damage and variation between shots are described. Transient expression of GUS occurred in pEmuGN-bombarded cells of nonregenerable suspension culture as well as in regenerable embryogenic callus of commercial sugarcane cultivar 463, and in suspension cultures capable of regeneration to plants. Parameters yielding transient GUS expression in up to 1055 cells per bombardment in homogeneous suspension cultures of sugarcane have been established with a mean of 206 expressing cells per bombardment over a series of 8 independent experiments. Approximately 4% of these transiently expressing cells continued to express GUS for extended periods, indicating probable stable transformation of intact cells of the commercial sugarcane cultivar. Microprojectile bombardment appears the most promising of the available gene transfer techniques for practical genetic transformation of sugarcane because most commercial cultivars readily form regenerable callus suitable for bombardment.