Stable Transformation of Plant Cells by Particle Bombardment/Biolistics
Particle bombardment, or biolistics, is a commonly used method for genetic transformation of plants and other organisms. Millions of DNA-coated metal particles are shot at target cells or tissues using a biolistic device or gene gun. The DNA elutes off the particles that lodge inside the cells, and a portion may be stably incorporated in the host chromosomes. A protocol for the generation of transgenic grapevines via biolistic transformation of embryogenic cell suspension cultures is detailed in this chapter. In a typical experiment, transient gene expression averaged nearly 8000 “hits” per bombarded plate. Five months after bombardment, there were nearly five putative transgenic embryos per bombarded plate. About half of the embryos were regenerated into confirmed transgenic plants. The basic bombardment procedures described are applicable to a wide range of plant genotypes, especially those for which embryogenic cell cultures are available. All users of particle bombardment technology will find numerous useful tips to maximize the success of transformation.
Key WordsBallistics biolistic biotechnology embryogenic cells gene gun genetic engineering grapevine microcarrier microparticle bombardment microprojectile bombardment particle acceleration particle bombardment particle gun plant transformation Vitis
- 8.Sanford, J. C., DeVit, M. J., Russell, J. A., et al. (1991) An improved, helium-driven biolistic device. Technique 3, 3–16.Google Scholar
- 16.Lloyd, G. and McCown, B. (1980) Commercially-feasible micropropagation of mountain laurel, Kalmia latifolia, by use of shoot-tip culture. Int. Plant Prop. Soc. Proc. 30, 421–427.Google Scholar
- 17.Russell, J. A., Roy, M. K., and Sanford, J. C. (1992) Major improvements in biolistic transformation of suspension-cultured tobacco cells. In Vitro Cell. Dev. Biol. 28P, 97–105.Google Scholar
- 19.Finer, J. J. and McMullen, M. D. (1991) Transformation of soybean via particle bombardment of embryogenic suspension culture tissue. In Vitro Cell. Dev. Biol. 27P, 17–182.Google Scholar