Nutritional and hormonal requirements of Ginkgo biloba embryo-derived callus and suspension cell culture

Abstract

Calli were obtained from Ginkgo biloba embryos grown on Murashige and Skoog (MS) medium. The G. biloba cells could grow on either MS or Gamborg B5 mineral salt medium supplemented with sucrose (3% and 2%, respectively) and naphthaleneacetic acid (NAA) and kinetin (K) in concentrations ranging from 0.1 to 2.0 mg·L⁻¹. Best growth and maintenance of callus cultures were achieved using MS medium supplemented with 2 mg·L⁻¹ NAA and 1 mg·L⁻¹ K (N₂K₁MS). Light was required to maintain healthy growth of the callus tissue.

In both MS and B5 based media, sucrose was hydrolyzed extracellularly before being taken up by Ginkgo cell suspension cultures. Specific growth rates of 0.13 d⁻¹ and 0.08 d⁻¹ were obtained in MS medium supplemented with 1 mg·L⁻¹ NAA, 0.1 mg·L⁻¹ K and 30 g·L⁻¹ sucrose (N₁K_0.1MS) and B5 medium supplemented with the same growth regulator regime and 20 g·L⁻¹ sucrose (N₁K_0.1B5) respectively. Complete phosphate and ammonium uptake was observed in 11 days when cultured in MS medium and 10 days and 4 days respectively when cultured in B5 medium. During the culture, G. biloba cells consumed only 64% and 29% of the nitrate content of N₁K_0.1MS and N₁K_0.1B5 media respectively. Maximum dry biomass concentrations were 13.4 g·L⁻¹ and 7.9 g·L⁻¹, and yields on carbohydrate were 0.39 and 0.45 in N₁K_0.1MS and N₁K_0.1B5 media respectively. The better performance of MS cultures came from the higher sucrose and nitrogen salts concentrations of this medium.