An approach to standardize transient expression of GUS in sesame (Sesamum indicum L.) using the Sonication-Assisted Agrobacterium tumefaciens-mediated Transformation (SAAT) method

摘要

Sonication-Assisted Agrobacterium-mediated Transformation (SAAT) dramatically improved Agrobacterium infection efficiency in Sesamum indicum (sesame) cotyledon. Different parameters of this method such as the duration of sonication, Agrobacterium concentration and acetosyringone concentration during co-cultivation was standardized to develop a balanced protocol that gave optimum beta-glucuronidase (GUS) expression with minimal tissue damage. Nine seconds was identified as the optimum duration for sonication. Agrobacterium concentration having OD600nm 1.0 was found not to induce tissue damage. Usage of 100 mu M acetosyringone, which facilitates T-DNA transfer by activating the vir gene in Agrobacterium, increased the transient GUS activity. The optimized protocol facilitated 85.56 +/- 0.35% transient GUS activity. Mimicking SAAT by increasing the concentration of acetosyringone during the co-cultivation period to 400 mu M in the conventional Agrobacterium-mediated transformation gave only 32.22 +/- 1.53% GUS activity. The results suggest that not only acetosyringone but also other compounds were released during formation of micro-wound by sonication that increased the infection efficiency.