目的:建立聚合酶链式反应-限制性片段长度多态性(PCR-RFLP)方法,用于湖北麦冬的分子鉴别。方法:在ITS 2区设计湖北麦冬、短葶山麦冬和麦冬的通用引物,经PCR反应后,对PCR产物进行限制性内切酶(Nae I)的酶切反应,酶切条件为37 ℃孵育60 min。结果:设计的通用引物对湖北麦冬和麦冬均能扩增出377 bp的单一条带,经酶切反应后,湖北麦冬被切为231 bp和146 bp 2条条带,麦冬仍为377 bp的单一条带,以此实现湖北麦冬的分子鉴别。结论:建立的PCR-RFLP方法能够实现湖北麦冬的准确鉴别,且该方法操作简单,特异性强,稳定性好,检测结果客观易判读。
Objective:To establish a polymerase chain reaction restriction fragment length polymorphism (PCR-RFLP) method for molecular identification of root of Liriope spicata. Methods: The universal primers for root of Liriope spicata, root of Liriope muscari and root of Ophiopogon japonicus were designed in ITS 2 region. After PCR reaction with the universal primers, the PCR products was digested by restriction endonuclease (Nae I) at 37 ℃ for 60 min. Results: Both root of Liriope spicata and root of Ophiopogon japonicus could be amplified by a single 377 bp band by the designed universal primers. The PCR chromatograms of root of Liriope spicata were cut into two bands, 231 bp and 146 bp after enzymatic digestion by Nae I, while root of Ophiopogon japonicus remained a single band of 377 bp, allowing molecular identification of root of Liriope spicata. Conclusion: Root of Liriope spicata can be identified accurately by the PCR-RFLP method. This method is simple, specific and stable, the identification results are objective and easy to interpret.
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