濒危南药白木香悬浮细胞体系的建立

Establishment of a cell suspension culture system of endangered Aquilaria sinensis(Lour.) Gilg

以白木香（Aquilaria sinensis）茎尖为材料,利用诱导形成的愈伤组织构建悬浮细胞体系。结果表明,诱导愈伤组织最适培养基为MS＋2.0 mg·L^-1 NAA＋1.0 mg·L^-1 6-BA;经12次继代培养的愈伤组织生长旺盛、质地疏松适于悬浮培养;将其置于液体培养基MS＋2.0 mg·L^-1 NAA＋1.0 mg·L^-1 6-BA＋500.0 mg·L^-1 CH中震荡培养,建成悬浮细胞体系。悬浮细胞生长曲线呈S型,初期增长缓慢,4~6天为对数增长期,7~12天进入平台期,12天以后细胞生长速度及活力迅速下降;GC-MS结果显示,培养0、3、6、9、12天悬浮细胞不含沉香倍半萜,使用100μmol·L^-1 MeJA处理24 h可检测到倍半萜。本实验构建的白木香悬浮细胞体系均一稳定、细胞活力良好,可作为研究伤害诱导白木香形成倍半萜的离体体系,同时也具备离体生产沉香倍半萜的潜力。

Aquilaria sinensis callus induced by stem tips were used to establish the suspension cell system. The results showed that the most suitable medium for callus induction and subculture is MS ＋ 2.0 mg·L^-1 NAA ＋ 1.0 mg·L^-1 6-BA. After 12 times of subculture, the energetic and loose callus, which were appropriate for cell suspension culture, were cultured and shook in liquid medium MS ＋ 2.0 mg·L^-1 NAA ＋ 1.0 mg·L^-1 6-BA ＋ 500.0 mg·L^-1 casein hydrolysate （CH） to establish the suspension cell system. The growth curve of suspension cells showed a ＂S＂ type. At the beginning of the culture, cell density increased slowly; during 4 to 6 days, suspension cells reached logarithmic growth period; during 7 to 12 days, suspension cells were in the platform period; butafter 12 days, cell density and activity went down obviously. Agarwood sesquiterpenes were not detected in the suspension ceils during the growth period, however, they could be detected in MeJA treated suspension cells. In this study, a stable and active growing suspension cell system was established, which was a proper system to study the mechanism of agarwood sesquiterpene formation, and additionally provided a potential way to generate agarwood sesquiterpenes through application of cell culture.