Li Hui, Qian Run, Tian Na, Li Yang-Hua, Jiang Chao, Yuan Yuan, Huang Lu-Qi
Aademician Workstation, Jiangxi University of Traditional Chinese Medicine Nanchang 330004, China.
School of Pharmacy, Anhui University of Chinese Medicine Hefei 230012, China.
Zhongguo Zhong Yao Za Zhi. 2020 Aug;45(15):3666-3671. doi: 10.19540/j.cnki.cjcmm.20200527.102.
Gastrodia elata is a kind of traditional Chinese medicinal materials and has good medicinal value. G. elata is divided into five varieties, which includes G. elata f. elata(proto variant), G. elata f. glauca, G. elata f. viridis, G. elata f. flavid and G. elata f. alba. Among them, G. elata f. elata and G. elata f. glauca have excellent characteristics and higher contents of gastrodin and polysaccharides. The hybrid of G. elata f. elata and G. elata f. glauca is present in markets, but the characteristics between hybrid and parent are not obvious and distinguished quickly and accurately. The aim of this study is to establish a PCR specific PCR identification method, which can identify G. elata f. elata, G. elata f. glauca and their hybrid. Based on the re-sequencing results of G. elata, we screened for the single nucleotide polymorphism(SNP) variation sites, and designed two pairs of specific primers(W291-F/W291-R and H255-F/H255-R). We further collected G. elata f. elata, G. elata f. glauca and their hybrid samples from different regions, established and optimized PCR method, and investigated and verified their tolerance and applicability. The results showed that when the annealing temperature was 48 ℃ and the number of cycles was 33, 255 bp specific band were obtained from G. elata f. glauca and hybrid by using specific primers W291-F/W291-R. When the annealing temperature was 51 ℃ and the number of cycles was 33, 291 bp specific band were obtained from G. elata f. elata and hybrid by using specific primers H255-F/H255-R. Our method could be used as a promising method to identify G. elata f. elata, G. elata f. glauca and their hybrid.
天麻是一种传统中药材，具有良好的药用价值。天麻分为五个变种，包括红天麻（原变种）、绿天麻、乌天麻、黄天麻和白天麻。其中，红天麻和绿天麻具有优良特性，天麻素和多糖含量较高。红天麻和绿天麻的杂交种已在市场上出现，但杂交种与亲本之间的特征不明显，难以快速准确区分。本研究的目的是建立一种PCR特异性鉴定方法，能够鉴定红天麻、绿天麻及其杂交种。基于天麻的重测序结果，筛选出单核苷酸多态性（SNP）变异位点，设计了两对特异性引物（W291-F/W291-R和H255-F/H255-R）。进一步收集不同地区的红天麻、绿天麻及其杂交种样本，建立并优化PCR方法，考察并验证其耐受性和适用性。结果表明，当退火温度为48℃、循环次数为33时，用特异性引物W291-F/W291-R可从绿天麻和杂交种中获得255 bp的特异性条带。当退火温度为51℃、循环次数为33时，用特异性引物H255-F/H255-R可从红天麻和杂交种中获得291 bp的特异性条带。本方法可作为一种有前景的方法用于鉴定红天麻、绿天麻及其杂交种。
