Utilization of sucrose during cocultivation positively affects Agrobacterium-mediated transformation efficiency in sugar beet (Beta vulgaris L.)

Abstract

Sugar beet (Beta vulgaris L.) is one of the most important industrial crops throughout world. With the availability of suitablegenetic transformation technologies, the yield, quality, and stress tolerance of sugar beet could be improved significantly. However, lowtransformation efficiencies seriously limit the application of molecular technologies to the genetic improvement of sugar beet. Withthe aim of improving gene transfer techniques for sugar beet, the effect of different sucrose concentrations during cocultivation onthe initial Agrobacterium-mediated transformation efficiencies in sugar beet was tested. To develop an efficient experimental systemthrough which the effect of sucrose could be tested, first, a prolific regeneration system was optimized by testing the effect of differentplant growth regulators on in vitro regeneration and rooting efficiencies from sugar beet cotyledonary node explants. The highestmean number of regenerated shoots per explant was obtained when the cotyledonary node explants excised from young seedlingswere grown on MS medium supplemented with 1.0 mg/L 6-benzylaminopurine. Using this regeneration system, the effect of differentconcentrations of sucrose included in the cocultivation medium on the initial genetic transformation efficiencies observed in T0 plantswas tested using an Agrobacterium tumefaciens strain carrying the pBin19/35S:GUS-INT construct. The inclusion of 4.5% sucrose inthe cocultivation medium resulted in significantly higher transformation (34.09%) and expression efficiencies (22.72%), confirmedby polymerase chain reaction and β-glucuronidase assays, respectively, in regenerated T0 seedlings. If translated into stably inheritedtransformation efficiencies, these findings could contribute to the success of genetic transformation studies in sugar beet and other cropsrecalcitrant to Agrobacterium-mediated transformation.