Abstract:

To establish a specific PCR method for rapid detection of plant origin components of bulbus Fritillariae cirrhosae, the HMGR gene sequence was downloaded from the NCBI database and compared. Degenerate primers were designed according to the homologous DNA fragments, and the genome from bulbus F. cirrhosae and other easily confused species were analyzed. Then specific primers were designed for longer specific nucleic acid fragment from bulbus F. cirrhosae. The total DNA was extracted from dry bulbs of bulbus F. cirrhosae. The specific primers BMH-TF/BMH-TR were designed and the PCR amplification conditions were optimized. Then the specificity and sensitivity were tested, and the PCR detection system specific for bulbus F. cirrhosae was successfully established. The PCR products were able to amplify a single band of 120 bp by agarose gel electrophoresis, and no amplification band was found in other samples and the blank under the same conditions. The sensitivity of this primer was 2 ng/μL, the lower limit of detection was 4 ng. The test results of 10 kinds of commercially available medicinal herbs proved that this method was simple, accurate and reliable with good reproducibility. This PCR method can identify bulbus F. cirrhosae rapidly, and has the advantages of rapidness and high specificity.

Key words: bulbus Fritillariae cirrhosae, multi-species origin, specificity, PCR, molecular identification