Induction and Suspension Culture of <i>Panax japonicus</i> Callus Tissue for the Production of Secondary Metabolic Active Substances

Abstract

Using Panax japonicus as research material, callus induction and culture were carried out, and high-yielding cell lines were screened to establish a suspension culture system that promotes callus growth and the accumulation of the “total saponins” (total content of triterpenoid glycosides or ginsenosides). Using the root as an explant, the medium for callus induction and proliferation was optimized by adjusting culture conditions (initial inoculation amount, carbon source, shaking speed, hormone concentration, culture time) and a high-yielding cell line with efficient proliferation and high total saponins content was screened out. The conditions of suspension culture were refined to find out the most suitable conditions for the suspension culture of callus, and finally, the suspension culture system was established. We found that the lowest (5%) contamination rate was achieved by disinfecting the fresh roots with 75% alcohol for 60 s, followed by soaking in 10% NaClO for 15 min. The highest induction rate (88.17%) of callus was obtained using the medium MS + 16.11 μmol·L−1 NAA + 13.32 μmol·L−1 6-BA + 30.0 g·L−1 sucrose + 7.5 g·L−1 agar. The callus was loose when the callus subcultured on the proliferation medium (MS + 5.37 μmol·L−1 NAA + 13.32 μmol·L−1 6-BA + 30.0 g·L−1 sucrose + 3.8 g·L−1 gellan gum) for 21 days. The callus growth was cultured in a liquid growth medium (MS + 5.37 μmol·L−1 NAA + 13.32 μmol·L−1 6-BA + 30.0 g·L−1 sucrose) with an initial inoculation amount of 40 g·L−1, a shaking speed of 110 r/min and darkness. Cell growth was fastest with a culture period of 21 days. We replaced the growth medium with the production medium (MS + 5.37 μmol·L−1 NAA + 13.32 μmol·L−1 6-BA + 30.0 g·L−1 glucose) for maximum accumulation of total saponins. [Conclusion] A callus induction and suspension culture system for the root of P. japonicus was established. In this way, we can promote the accumulation of total saponins in callus cells and provide a basis for large-scale cell culture and industrial production of medicinal total saponins.

Keywords

• <i>Panax japonicus</i>
• callus tissue
• culture time
• suspension culture
• “total saponins” triterpenod glycosides
• ginsenosides