Optimization of Transformation Efficiency of Suspension Cultured Vitis vinifera cv. Chardonnay Embryogenic Cells

Abstract

Vitis vinifera cv. Chardonnay suspension cultures were established from proembryogenic mass and employed for optimizing Agrobacterium-mediated transformation system. One-factor-at-a-time experiment revealed that OD600 of Agrobacterium, time of inoculation, co-cultivation, and cell-drying before inoculation significantly affected the transformation efficiency which reached maximum 21.5% at the following conditions: 0.8 of OD600, 25 min of inoculation, 2 d of co-cultivation, and 10 min of cell drying. Response surface methodology experiments based on a five-level, four-factor central-composite rotatable design were then used to optimize these selected factors. The optimized conditions for Chardonnay grape transformation were: 0.8711 of OD600, 28.9 min of inoculation, 2.25 d of co-cultivation and 11.76 min of cell drying. After optimization, transformation efficiency was 26.2% and there were no interactions among different factors.

Keywords

• transformation
• Chardonnay
• suspension cultures
• response surface methodology