EXTRACELLULAR ENZYMES OF ERWINIA-CAROTOVORA ELIMINATE THE NEED FOR AZACYTIDINE TREATMENT FOR HIGH-FREQUENCY TRANSFORMATION OF ARABIDOPSIS-THALIANA

This paper reports a part of our studies on large-scale T-DNA-mediated gene tagging in Arabidopsis thaliana. To enhance the chance of taggin g specific stress-responsive genes of this species by monitoring the p referential insertion of the T-DNA into the actively transcribed loci, we exposed the root explants to low temperature (LT), abscisic acid ( ABA), and extracellular enzymes (EXE) of the plant pathogen Erwinia ca rotovora prior to transformation by Agrobacterium tumefaciens. Both LT and ABA reduced the frequency of transformation; with these treatment s, the average transformation frequencies were 8.1% and 2.6%, respecti vely. However, in explants pretreated with EXE the transformation freq uency was 89.0%, similar to that obtained in control materials (92.6%) . Transgenic calli developed from these explants did not require any t reatment with azacytidine (azaC) for efficient shoot regeneration. Fur thermore, this treatment enhanced multiple insertion of the T-DNA into the plant genome; within a population of EXE-treated transgenic plant s, the number of lines harboring at least three copies of the integrat ed T-DNA was much higher (61%) than that observed in an untreated popu lation (34%).