目的 利用细胞悬浮培养技术生产霍山石斛单细胞，解决石斛药用植物资源短缺问题。方法 以霍山石斛叶片等外植体为实验材料，通过不同的制备方法获得霍山石斛单细胞，研究霍山石斛细胞培养的多种影响因素，应用正交设计优化霍山石斛细胞悬浮培养的条件。结果 在蔗糖35 g/L，NH₄NO₃ 1.3 g/L，pH 5.6，激素IAA 0.4 mg/L，6-BA 1.0 mg/L，培养时间14～18 d，在此条件下培养的霍山石斛细胞数为9.42×10⁵个/mL。结论 经过4次继代培养后石斛细胞多为圆状或椭圆状，细胞碎片减少。应用优化的细胞悬浮培养条件，能获得较大量的霍山石斛细胞。

Objective Single-cell of Dendrobium huoshanenes was cultured by using cell suspension culture for solving the problem of resource shortage of medicinal plant D. huoshanenes. Methods The single-cell of D. huoshanenes was obtained by different preparing ways, effects of various factors on cell growth were studied and the optimal growing conditions of cell suspension culture were investigated by orthogonal design. Results Through the orthogonal test, the results indicated that the optimum conditions for suspension culture of D. huoshanenes cell were as follow: is that the concentration of sucrose was 35 g/L , NH₄NO₃ was 1.3 g/L, pH value was 5.6, the hormone IAA was 0.4 mg/L , 6-BA was 1.0 mg/L, the best culture time was 14－18 d. Under optimal culture conditions, the concnentration of single cells in this test was 9.42×105 cell/mL. Conclusion After four successive subculture, cellular morphology of D. huoshanenes shows circular shape or elliptic shape and the chip of cell is decreased. The higher concentration of D. huoshanenes cell could be obtained by using the optimal growing conditions of cell suspension culture.

安徽省高校自然科学基金研究项目(KJ2009B244Z)