Transformation of crown gall resistant and susceptible <i>Vitis</i> genotypes by <i>Agrobacterium vitis</i>

Abstract

Transformation of crown gall-susceptible and -resistant Vitis genotypes by Agrobacterium vitis strain CG49 was studied using uidA (GUS) in the p35SGUSINT construct. When greenhouse-grown material propagated through tissue culture was inoculated with CG49(p35SGUSINT) in vitro, the highly crown gall-susceptible V. vinifera Cabernet Sauvignon displayed GUS activity on 53 % of inoculated explants vs. 5 % for the resistant V. amurensis and 0 % for the resistant Couderc 3309. Response of Cabernet Sauvignon suggested a strong effect of shoot polarity on transformation. Inoculation of basal vs. apical explant surface in Cabernet Sauvignon indicated transformation in 88 % of basal inoculated explants with no transformation from apical inoculation. Basal inoculations indicated no transformation of V. amurensis and transformation in 10 % of Couderc 3309 explants. Inoculation of intact plants with CG49(p35SGUSINT) produced GUS-positive sites at 56 % of inoculated sites in Cabernet Sauvignon, 10 % of V. amurensis inoculated sites and 9 % of Couderc 3309 inoculated sites. Resistance to crown gall in these genotypes appears to be due to reduced susceptibility to transformation by A. vitis rather than post-transformation phenomena. These studies were complicated by production of GUS-positive spots from in vitro inoculations using wild-type CG49. Resident microorganisms producing b-glucuronidase may proliferate after tissue degradation by A. vitis-induced cell disruption. Use of in vitro internodal explants from tissue culture-propagated vines greatly reduced GUS expression from control CG49 inoculations and these were readily distinguished (by appearance and location) from GUS-positive spots resulting from transformation with uidA.