摘要
目的构建适合铁棒锤种子蛋白质研究的聚丙烯酰胺凝胶电泳(SDS-PAGE)体系,为铁棒锤种子纯度检测及鉴别研究提供技术支撑。方法采用6种蛋白质提取法提取伏毛铁棒锤种子蛋白质,并筛选出最佳提取方法,在此基础上研究不同分离胶浓度、料液比、上样量对伏毛铁棒锤种子蛋白电泳分离效果的影响,并采用筛选出的蛋白质最佳提取分离方法对铁棒锤和伏毛铁棒锤种子进行鉴定。结果巯基乙醇法提取的伏毛铁棒锤种子蛋白质电泳图谱效果较佳,电泳谱带数最多(14条)、最清晰,适合于其种子纯度检测;分离胶浓度为15%、料液比为1∶10(M/V)、蛋白上样量稀释5倍时,其种子蛋白质电泳分离效果较好;铁棒锤和伏毛铁棒锤种子蛋白质电泳图谱在蛋白质谱带位置和强弱上有一定差异。结论本研究建立的铁棒锤种子蛋白样品制备方法及SDS-PAGE技术可用于铁棒锤种子纯度检测,其蛋白质图谱可作为铁棒锤与伏毛铁棒锤种子的鉴别依据。
Objective To establish the appropriate SDS-PAGE system for study on protein from Aconitum pendulum Busch. seeds;To provide technical support for the purity detection and identification of Aconitum pendulum Busch. seeds. Methods Six protein extraction methods for Aconitum pendulum Busch. seeds were tested, and the best extraction method was selected out. Based on the optimal extraction method, the effects of different separation gel concentrations, solid-liquid ratios and sample loadings on the electrophoretic separation of protein from Aconitum pendulum Busch. seeds were studied. The optimal protein extraction and separation method was used to identify seeds of Aconitum pendulum Busch. and Aconitum flavum Hand. Mazz. Results The electrophoresis pattern of Aconitum flavum Hand. Mazz. extracted by thiol ethanol method was better, and the electrophoresis band number was the most (14) and the clearest, which were suitable for its seed purity detection. When the concentration of separation gel was 15%, the ratio of material to liquid was 1:10 (M/V), and the amount of protein sample was diluted by 5 times, the separation effect of seed protein was better. The electrophoresis patterns of Aconitum pendulum Busch. and Aconitum flavum Hand. Mazz. had some differences in the position and strength of the protein bands. Conclusion The developed methods of seed protein extracting and SDS-PAGE electrophoresis technique can be used for seed purity detection, and the protein fingerprints can be applied to identify seeds of Aconitum pendulum Busch. and Aconitum flavum Hand. Mazz.
作者
陆国弟
杨扶德
陈红刚
王惠珍
杜弢
邢豪
LU Guodi;YANG Fude;CHEN Honggang;WANG Huizhen;DU Tao;XING Hao(Department of Pharmacy,Gansu University of Chinese Medicine,Lanzhou 730000,China)
出处
《中国中医药信息杂志》
CAS
CSCD
2019年第6期64-69,共6页
Chinese Journal of Information on Traditional Chinese Medicine
基金
国家现代农业中药材产业技术体系建设专项基金(CARS-21)
甘肃省高校中(藏)药化学与质量研究省级重点实验室开放基金(zzy-2016-04)
甘肃省中药质量与标准研究重点实验室培育基地开放基金(ZYZL16-003)
兰州市科技计划项目(2016-3-116)
关键词
铁棒锤
种子
聚丙烯酰胺凝胶电泳
蛋白质提取
鉴定
Aconitum pendulum Busch.
seed
SDS-PAGE
extraction of protein
identification
