利用正交设计优化金钗石斛的转化体系
Optimization of Agrobacterium-Mediated Transformation System for Dendrobium nobile Lindl by Orthogonal Design
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摘要: 为优化金钗石斛的转化体系,以其类原球茎(PLBs)为材料,利用正交设计研究了影响农杆菌介导转化的因素.结果表明:PLBs预培养1 d、侵染的菌液OD600值1.2、乙酰丁香酮(AS)100μmol/L、共培养5 d时,每100mg鲜物质量材料上的GUS染色蓝点数最多;单因子试验表明AS适宜的浓度为50μmol/L;PCR结果显示得到了转化呈阳性的PLBs;初步表明此体系可用于金钗石斛的遗传转化研究.
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关键词:
- 金钗石斛,农杆菌,转化,正交设计
Abstract: In order to optimize the Agrobacterium-mediated transformation system for Dendrobium nobile Lindl, effects of preculture time, bacterial concentration, acetosyringon (AS) concentration and co-cultivation time on Agrobacterium-mediated transformation of protocorm like bodies (PLBs) of D. nobile Lindl were studied in an experiment with orthogonal design. The results showed that number of GUS staining blue spots per 100 mg flesh weight of PLBs was the greatest in the treatment of the combination of preculture for one day, 1.2 of bacterial concentration (OD600), 100 μmol/L of concentration of AS and co-culture for five days. Single factor experiment showed that the appropriate concentration of AS was 50 μmol/LPCR confirmed that positive transformed PLBs were obtained. It is tentatively concluded that this system can be used in the study of transformation of D. nobile Lindl. -
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