PCR-RFLP法鉴别罗布麻与混淆品白麻及其psbA-trnH序列分析  

Distinguishing Apocynum venetum from its Confusion Apocynum pictum with PCR-RFLP and Sequence Analysis of PsbA-trnH

作　　者：刘流 邰顺章 支荣荣 LIU Liu;TAI Shunzhang;ZHI Rongrong(Yancheng Food and Drug Supervision and Testing Center,Yancheng 224055,China)

机构地区：[1]盐城市食品药品监督检验中心,盐城224055

出　　处：《药学与临床研究》2021年第6期411-415,共5页Pharmaceutical and Clinical Research

摘　　要：目的:基于叶绿体psbA-trnH基因间隔区序列,建立一种PCR-RFLP(聚合酶链式反应-限制性片段长度多态性)方法鉴别罗布麻(Apocynum venetum L.)及其混淆品白麻(Apocynum pictum),并验证psbA-trnH序列酶切位点种间特异性和种内保守性。方法:将罗布麻和白麻提取总DNA,扩增psbA-trnH序列,扩增产物双向测序。序列分析发现,罗布麻含有特异性酶切位点SspⅠ。结果:扩增产物psbA-trnH为300~400bp。罗布麻psbA-trnH产物均可以被SspⅠ酶切成两条条带,白麻却不能被SspⅠ酶切。生物信息分析结果表明,在罗布麻属中,罗布麻psbA-trnH序列的SspⅠ酶切位点具有种间特异性和种内保守性,在夹竹桃族进化树中,罗布麻与白麻距离最近。结论:本实验建立的PCR-RFLP方法可以有效鉴别罗布麻和白麻。Objective:To establish a method for distinguishing Apocynum venetum from its confusion Apocynum pictum with PCR-RFLP(Polymerase chain reaction-Restriction Fragment Length Polymorphism)based on chloroplast intergenic psbA-trnH spacer sequence.To verify if the restriction enzyme site is specific among species of genus Apocynum and conserved in Apocynum venetum.Methods:After extracting total DNA,amplifying psbA-trnH region and paired end sequencing,sequence analysis was conducted.Results:The psbA-trnH PCR products were between 300-400 bp.There was a specific restriction enzyme cutting site SspⅠin the Apocynum venetum psbA-trnH region.Apocynum venetum psbA-trnH PCR products were all cut into two parts with Ssp I,Apocynum pictum psbA-trnH PCR products could not be cut apart with SspⅠ.Bioinformatic analyzing showed that SspⅠsite in the psbA-trnH sequence of Apocynum venetum was specific among species of genus Apocynum and conserved in Apocynum venetum.Apocynum venetum was the closest to Apocynum venetum in the Trib.Apocyneae Airy Shaw phylogenetic tree.Conclusion:This PCR-RFLP method can efficiently distinguish Apocynum venetum from Apocynum pictum.

关 键 词：罗布麻 白麻 聚合酶链式反应-限制性片段长度多态性 PSBA-TRNH 分子鉴定 酶切位点特异性 

分 类 号：R927[医药卫生—药学] Q789[生物学—分子生物学]