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目的:利用杜仲药材及其混伪品的rDNA-ITS序列差异设计DNA特异引物,实现杜仲药材的快速分子标记鉴定。方法:从NCBI数据库下载杜仲药材及其混伪品的rDNA-ITS序列,比对获得差异DNA片段,设计特异性引物,优化PCR扩增条件和检测方法。结果:扩增产物经琼脂糖凝胶电泳和荧光反应检测,杜仲药材能扩增400bp的鉴定条带,加入SYBR GreenⅠ染料后在紫外光下有绿色荧光,而其混伪品不具特异条带和绿色荧光。结论:特异性PCR能有效鉴别杜仲药材及其混伪品,通过条件优化显著缩短PCR扩增时间,荧光染料检测能更直观显示鉴定结果,为杜仲药材的快速鉴定提供了新的思路和方法。
OBJECTIVE: To design DNA-specific primers based on the rDNA-ITS sequence differences of Eucommia ulmoides medicinal herbs and their adulterants, and to achieve rapid molecular marker identification of Eucommia ulmoides herbs. Methods: The rDNA-ITS sequences of Eucommia ulmoides Oliv. And its adulterants were downloaded from the NCBI database, and different DNA fragments were obtained by comparison. The specific primers were designed and the PCR amplification conditions and detection methods were optimized. Results: The amplified product was detected by agarose gel electrophoresis and fluorescence reaction. The Eucommia ulmoides medicinalum could amplify the 400bp identification band. After addition of SYBR GreenⅠ dye, there was green fluorescence under UV light, Green fluorescence. Conclusion: Specific PCR can effectively distinguish Eucommia ulmoides and its adulterants, and can significantly shorten the PCR amplification time by optimizing the conditions. Fluorescent dye detection can more directly show the identification results, which provides a new idea and method for the rapid identification of Eucommia ulmoides medicinal materials.