该文选择8种含有地格达类蒙药原料药材的蒙成药为对象,首先对基原植物肋柱花和苦地丁样品采用改良CTAB法提取其总DNA,使用psb A-trn H片段进行扩增、测序,与Gen Bank下载的8种蒙成药中其他原料药材psb A-trn H序列进行同源比对后根据其变异位点设计特异性鉴别引物。同时对8种蒙成药进行总DNA的提取,并使用DNA纯化试剂盒进行纯化,通过PCR反应对叶绿体rbc L序列进行扩增,再分别选择肋柱花和苦地丁鉴别引物进行扩增,并将扩增后的产物进行测序。所得序列校正、比对后,进行比对分析。结果表明,地格达-4汤、地格达-8散、地格达-4散中均含有原料药材肋柱花,苦地丁特异鉴别引物扩增分析可以鉴定出利胆八味散、伊赫哈日-12和阿嘎日-35中含有苦地丁。该研究结果说明,位点特异PCR方法用于鉴定蒙成药中原料药材具有一定的可行性。
To explore a new method for identification of Mongolian patent medicine （MPM） by PCR amplification of specific al- leles. Eight kinds of MPM were used to study the identification of ＂Digeda＂ raw materials. The total DNA of Loma~gonium rotatum and Corydalis bungeana samples were extracted through modified CTAB method, psbA-trnH sequence was amplified by PCR and sequenced directionally. Specific primer was designed. The DNA of 8 kinds of MPM also was extracted and purified by the commercial DNA puri- fication kits. The rbcL and two pair of specific primers sequences were amplified. The specific amplified products were sequenced in forward directions. All specific sequences were aligned and were analyzed. The results indicated that L. rotatum can be identified by specific primers from Digeda4 Tang, Digeda-8 San, Digeda4 San, and C. bungeana medicinal materials can be identified by specific primers from Li Dan Ba Wei San, Yi He Ha Ri-12 and A Ga Ri-35. PCR amplification of specific alleles can stably and accurately dis- tingnish raw medicinal materials in MPM.
China Journal of Chinese Materia Medica
Mongolian patent medicine
＂Digeda＂ medicinal plants
PCR amplification of specific allele