不同花色白芨四种DNA条形码的序列分析  

Sequence Analysis of Different Flower Color of Bletilla striata Based on Four DNA Barcoding Markers

作　　者：曾丽娜 刘显义 石建龙 梁大勇 张洁 张婷婷 任建国[1,3] 刘红美 ZENG Li-na;LIU Xian-yi;SHI Jian-long;LIANG Da-yong;ZHANG Jie;ZHANG Ting-ting;REN Jian-guo;LIU Hong-mei(a.Sehool of Biology and Engineering;1b.Engineering Researeh Center of Medieal Bioteehnology,Guizhou Medieal University,Guiyang 550025,China;2.Zunyi Longehi Biologieal Teehnology Co.,Ltd.,Zunyi 563100,Guizhou,China)

机构地区：[1]贵州医科大学生物与工程学院,贵阳550025 [2]贵州省遵义市龙驰生物科技有限公司,贵州遵义563100 [3]贵州医科大学医药生物技术工程研究中心,贵阳550025

出　　处：《湖北农业科学》2018年第17期106-110,共5页Hubei Agricultural Sciences

基　　金：贵州省重点学科(培育)项目(ZDXK[2016]26号);贵州医科大学校级重点实验室建设项目(校重点实验室2016002);贵州医科大学校级工程研究中心建设项目(校工程中心2016002);贵州省细胞与基因工程创新群体重大研究项目(黔教合KY字[2016]031)

摘　　要：采用CTAB法提取2种花色白芨基因组DNA,选用trnH-psbA、rbcL、ITS、LSU D1-D3 4种植物DNA条形码通用引物进行PCR扩增,分析黄花、紫花白芨(Bletilla striata)4种DNA条形码的序列,扩增产物纯化后连接T载体并测序,用DNAMAN软件对测序结果进行多序列比对分析。再根据SNP位点设计引物,利用位点特异性PCR扩增进行鉴定。结果表明,4对植物DNA条形码通用引物均能进行扩增,其中trnH-psbA序列没有发现SNP位点、rbcL序列出现2个SNP位点、ITS序列出现62个SNP位点、LSU D1-D3序列出现3个SNP位点。根据LSU D1-D3序列上的2个SNP位点设计的引物DUAN5(5′-TAGTAACGGCGAGCGAAC-3′)和JDHH(5′-TTATCAGCTTCCTTGCGCCCG-3′)能准确鉴定出黄花白芨,成功获得鉴定黄花白芨的引物序列。GenomicDNA was extracted from the two kinds of color of Bletilla striata by using the CTAB method. To analysis the sequence of different flower color of Bletilla striata based on the four kinds of DNA barcode for further identification.Four DNA barcoding markers（trnH-psbA,rbcL,ITS and LSU D1-D3） were selected for PCR. PCR product was ligated into T vector for sequencing. The sequencing results were analyzed by using DNAMAN software. According to the SNP loci of different color of Bletilla striata, the primers were designed for identifying different color of Bletilla striata. Result showed that：two kinds of flower color of Bletilla striata genomicDNA can be amplified based on 4 pairs primers of plant DNA barcode markers and SNP loci were found. None SNP locus was found in trnH-psbA sequence. Two SNP loci were found in rbcL sequence. Sixty-two SNP loci were found in ITS sequence. Three SNP loci were found in LSU D1-D3 sequence. Primer DUAN5（5′-TAGTAACGGCGAGCGAAC-3′） and JDHH（5′-TTATCAGCTTCCTTGCGCCCG-3′） can be used for identifying the yellow flower color of Bletilla striata according to the SNP loci in LSU D1-D3 sequence. The primers were found for identifying the yellow flower color of Bletilla striata.

关 键 词：白芨(Bletilla striata) DNA条形码 PCR SNP 

分 类 号：R282[医药卫生—中药学]