GENETIC CHANGES INDUCED IN HIGHER PLANTS BY A UV LASER MICROBEAM

Abstract

Plant cell walls can be perforated by a UV laser microbeam. Cells and immature pollen grains (microspores) of Brassica napus L. continued to grow in culture after being punctured by a laser pulse. During laser treatment, uptake of buffer into the cytoplasm of cells could be achieved by plasmolyzing cells in a hypertonic solution. Using this approach, plasmid DNA carrying a gene for bacterial glucuronidase was introduced into B. napus cells. The marker gene was expressed by 70% of the treated cells. Likewise, the gene for resistance to the antibiotic hygromycin B was introduced into B. napus cells and was stably integrated into growing cells. Furthermore, DNA was also incorporated into chloroplasts. First, DNA was injected into the cytoplasm and then the organelles were punctured by laser pulses. When a marker gene coding for triazine resistance was introduced into the organelles, transient expression at the level of callus cultures was observed.