摘要
目的采用随机扩增DNA多态性(RAPD)技术扩增鹿茸的线粒体DNA,扩增产物用无胶筛分毛细管电泳紫外检测,从而建立鹿茸的高效毛细管电泳-随机扩增DNA多态性(HPCE-RAPD)指纹图谱。方法对影响聚合酶链式反应(PCR)扩增和毛细管电泳的因素进行优化,在20 mmol·L-1NaH2PO4-Na2HPO4-2 mmol·L-1EDTA缓冲溶液[0.8%(W/V)HPMC,15 mmol·L-1TBAP,pH 7.3]、进样电压-10 kV、分离电压-8 kV的优化条件下,对梅花鹿茸、马鹿茸、驯鹿茸以及人工伪鹿茸进行分析。结果将获得的指纹图谱进行相似度分析,发现不同鹿茸样品的指纹图谱相似度有明显区别,可用于鹿茸样品的快速真伪鉴别。结论 HPCE-RAPD指纹图谱结合了RAPD技术多态性丰富、检出率高、技术简便和毛细管电泳高效、快速、灵敏等优点,适用于鹿茸及其伪品的快速鉴别,通过进一步研究,本方法还可广泛应用于其他易混中药材的快速鉴别。
OBJECTIVE To establish a high performance capillary electrophoresis-random amplified polymorphic DNA （HPCE-RAPD） fingerprinting method of antler. METHODS Mitochondrial DNA of Cornu Cervi Pantotrichum was amplified with RAPI） technique and the PCR products were detected with non-gel sieving capillary electrophoresis combined with UV detection. Factors that could affect the PCR amplification and capillary electrophoresis were optimized as follows： 20 mmol · L^-1 NaH2PO4-Na2HP4-2 mmol· L^-1 EDTA buffer solution [0. 8% （W/V） HPMC, 15 mmol· L^-1 TBAP and pH 7. 3）, - 10 kV injection vohage and -8 kV separation voltage. RESULTS The sim- ilarity analysis of obtained fingerprints showed that the fingerprints of different antlers were significantly different in the similarity. CONCLUSION HPCE-RAPD fingerprinting technique shows some advantages in the identification of Cornu Cervi Pantotrichum samples, such as rich polymorphism, high detection rate, convenience, efficiency, rapidness and sensitivity, indicating that it would be suitable for the quick identification of the authenticity of Cornu Cervi Pantotrichum samples. It is believed that this approach can be widely applied for the rapid identification of other easily confused Chinese medicinal materials if it is improved through the further investigation.
出处
《中国药学杂志》
CAS
CSCD
北大核心
2014年第15期1300-1305,共6页
Chinese Pharmaceutical Journal
基金
吉林省科技发展计划项目(20090906)
吉林省战略性新兴产业和高新技术产业发展专项(2013G030)
吉林省教育厅"十二五"科学技术研究项目(吉教科合字2014第212号)资助
关键词
鹿茸
无胶筛分毛细管电泳
随机扩增DNA多态性(RAPD)
指纹图谱
Cornu Cervi Pantotrichum
non-gel sieving capillary electrophoresis
random amplified polymorphic DNA （RAPD）
fingerprint
作者简介
孙绩岩，女，硕士研究生研究方向：中药DNA指纹技术与应用.
通讯作者：苑广信，男，博士，讲师研究方向：中药质量控制方法研究Tel／Fax：（0432）64608281E-mail：yuanguangxin2007@163．com
