Optimization of the uidA gene transfer of Rosa hybrida via Agrobacterium tumefaciens: an assessment of factors influencing the efficiency of gene transfer

Abstract

To develop a transformation protocol of Rosa hybrida ‘Samantha’ via Agrobacterium tumefaciens, the authors examined the effect of different factors on T-DNA transfer by measuring transient expression levels of an intron-containing β-glucuronidase gene. The results indicate that explant, light condition, salt concentration and acetosyringone (AS) concentration in co-culture medium are the most important factors, and factors like co-culture temperature, co-culture period and bacteria density have a strong effect on the growth of bacteria and then T-DNA transfer. Optimized co-cultivation was performed by inoculation of embryogenic callus with bacteria at a density of OD600=0.5–0.8 for 20 min and co-culture in darkness under 23°C on medium with 1/2 MS salts and 300 µmol·L−1 AS for 3 d.