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目的探讨特异性PCR方法在鉴定天花粉中的应用价值。方法选取16份天花粉正品和7份天花粉易混淆品,提取基因组DNA,按照天花粉核糖体RNA中ITS保守区域设计的3个引物进行PCR扩增,1.5%琼脂糖凝胶电泳检测PCR产物。结果引物P1(5’-TTCGTTTGAGTTCCTTGGCG-3’)扩增效果最好,最佳退火温度为45℃;560bp和960bp条带为天花粉特征鉴定条带,不同栝楼亚种的其他条带略有不同,可将其作为栝楼鉴定的辅助条带。结论锚定引物扩增多态性DNA技术(APAPD)是一种极有发展前景的中药材分子鉴定方法,可为中药材DNA指纹图谱提供更多信息。
Objective To investigate the value of specific PCR in the identification of TCS. Methods Genomic DNA was extracted from 16 genuine TCS and 7 TCS. Three primers were designed according to ITS conserved region of the microsomal ribosomal RNA. The PCR products were detected by 1.5% agarose gel electrophoresis. Results The primers P1 (5’-TTCGTTTGAGTTCCTTGGCG-3 ’) had the best amplification efficiency and the best annealing temperature was 45 ℃. The bands of 560bp and 960bp belonged to TCS bands, and the other bands of Brachypodium distichum were slightly different , Can be used as an identification of Qionglai supporting strip. Conclusion APAPD is an extremely promising method for molecular identification of Chinese herbal medicines and can provide more information on DNA fingerprinting of Chinese herbal medicines.