中药材五灵脂特异性PCR鉴别方法的研究

Specific PCR Identification of Faeces Trogopterori

目的建立快速检测五灵脂中药材动物源性成分的特异性PCR法。方法从NCBI数据库下载复齿鼯鼠及相关混伪品的16S rDNA序列,通过比对获得特异性DNA片段,设计特异引物,从复齿鼯鼠的干燥粪便中提取DNA,优化PCR扩增条件,通过特异性、灵敏度实验验证,建立特异性PCR检测体系。结果 PCR产物经琼脂糖凝胶电泳后扩增出一条340 bp左右条带,而其他样品在相同条件下无扩增条带,该引物的灵敏度可达4.43ng/μl。对15种市售药材的检测结果证明该方法简便可行。结论此PCR法可快速鉴别五灵脂,不受材料和干燥程度的影响,具有操作简便、快速、特异性高的优点。

Objective To establish a specifc PCR method for rapid detection of animal origin components of Faeces trogopterori. Methods The 16S rDNA sequence was downloaded from the NCBI database and the specific DNA fragments were obtained by comparison. The specifc primers were designed and the DNA was extracted from the dry stool of Trogopterus xanthipes. The PCR amplification conditions were optimized, through the specificity and sensitivity experiments, specific PCR detection system was established. Results The PCR products were able to amplify a single band of about 340 bp by agarose gel electrophoresis. However, no amplifcation bands were found in other samples under the same conditions. The sensitivity of this primer was 4.43 ng/μl. The test results of 15 kinds of commercially available medicinal herbs prove that this method is simple and feasible. Conclusion The PCR method can identify Faeces trogopterori rapidly, with independence of material and degree of dryness, as well as the advantages of simpleness, rapidness and high specifcity.