Development and Evaluation of Antler DNA Detection Kit
- VernacularTitle: 鹿茸DNA检测试剂盒的研制与评价
- Author:
Yan XU
1
Author Information
- Publication Type:Journal Article
- Keywords: Antler; Evaluation; Kit; Molecular biology
- From: Chinese Pharmaceutical Journal 2020;55(11):889-894
- CountryChina
- Language:Chinese
- Abstract: OBJECTIVE: To study the DNA fingerprint characteristics of the precious Chinese medicinal antler, identify and analyze the velvet antler at the molecular level, and develop the velvet DNA detection kit and evaluate its performance. METHODS: Using antler cytochrome C oxidase subunit Ⅰ (Co Ⅰ) and velvet mitochondrial cytochrome b (cytochrome b, Cyt b) as target genes, five pairs of specific primers were designed by bioinformatics technology, and the oxidase subunit Ⅰ of cytochrome C, mt DNA CoⅠ 1 (NC_013834.1), was determined to be the best specific primer by experimental screening. Polymerase chain reaction (PCR) amplification was conducted on velvet antler samples, and the PCR reaction system and reaction conditions were optimized to develop a velvet DNA detection kit. The performance of the kit was evaluated and commercially available antler samples were tested. RESULTS: The purity of the DNA extracted by the antler DNA test kit was 1.785-1.906. The specificity of the kit was reflected in the detection of genuine bands, and there was no band in the fakes. The sensitivity was up to 3.125 ng•μL-1. The kit was still effective after 5, 10, 15 and 20 freeze-thaw cycles. The results of repeated detection of positive and negative products for three times were consistent. The kit could be stored for up to one year at -20 ℃. The quality analysis was carried out on 12 commercial samples in five regions, and the pass rate was 66.67%. CONCLUSION: The velvet DNA detection kit developed by our team has strong specificity, high sensitivity, good stability and repeatability, with simple operation, small sample volume, wide application range, low cost and accurate results. It provides a scientific, accurate and reliable identification method for Chinese medicinal materials velvet products. At the same time, it also shows that the quality of commercially available velvet samples is uneven, and there are many adulterants.
