基于DNA条形码psbA-trnH,matK,trnL对不同地理居群野菊和药用菊的鉴定研究  

Identification of Chrysanthemum indicum in different geographical populations and Ch. morifolium based on DNA barcodes of psbA-trnH,matK and trnL

作　　者：陈芙蓉 汪涛[1] 郭巧生[1] 朱再标[1] 杨锋 邹庆军 张亚静 CHEN Fu-rong;WANG Tao;GUO Qiao-sheng;ZHU Zai-biao;YANG Feng;ZOU Qin-jun;ZHANG Ya-jing(Institute of Chinese Medicinal Materials , Nanjing Agricultural University, Nanjing 210095, China)

机构地区：[1]南京农业大学中药材研究所,江苏南京210095

出　　处：《中国中药杂志》2019年第4期660-665,共6页China Journal of Chinese Materia Medica

基　　金：国家自然科学基金青年基金项目(81503180);中央高校基本科研业务费专项资金项目(KJQN201643;KYZ201608)

摘　　要：利用DNA条形码技术通过psbA-trnH,mat K,trn L序列分析,建立一种快速鉴定野菊和药用菊的方法。提取21份样品的基因组DNA,对psbA-trnH,mat K,trn L片段进行PCR扩增,双向测序,获得其psbA-trnH,mat K,trn L序列信息;运用MEGA 7. 0对所有样品的psbA-trnH,mat K,trn L序列共63条进行比对、计算种内种间遗传距离,分析psbA-trnH+mat K+trn L组合序列SNPs分布,构建Neighbor-Joining(NJ)系统进化树。结果显示,野菊,野菊(菊花脑)和药用菊的种内种间遗传距离重合; psbA-trnH+mat K+trn L组合序列的SNPs分析显示其组合序列共有19个核苷酸多态性位点(SNPs),野菊较野菊(菊花脑)、药用菊呈现更为明显的序列多态性,psbA-trnH序列表现出较为明显的长度变异; NJ树显示药用菊与野菊、野菊(菊花脑)区别开来,其中编号为C2～C5的药用菊单独聚为一亚支,置信度为62%,编号为Z9,Z10的野菊均采自甘肃省,单独聚为一支,置信度为77%;同时也发现来自西北地域的野菊样品,其psbA-trnH和trn L序列信息变化与地域和环境之间有明显的相关,野菊与野菊(菊花脑)具有明显的分化,同时还是药用菊的进化来源。因此,将psbA-trnH+mat K+trn L组合序列作为DNA条形码能准确、快速的鉴别野菊和药用菊,为其种质资源鉴定及物种鉴别提供了重要依据。DNA barcode technology was used to establish a rapid identification method of Chrysanthemum indicum and Ch. morifolium based on psbA-trnH, matK and trnL sequences. The total DNA was extracted from 21 samples collected, and the psbA-tmH, matK, trnL sequences were amplified by PCR and sequenced. The information of these sequences were obtained. We aligned all 63 sequences, calculated the intraspecific and interspecific distances, analysed the SNPs distribution of psbA-trmH+matK+trnL coml)ination sequences and constructed the Neighbor-joining( NJ) Tree, using MEGA 7. 0. The results showed that the genetic distances of Ch. irulicum, Ch. indicum (Juhuanao) and Ch. morifolium were overlapped. Tlie SNPs analysis of psbA-trnH+matK+trnL combination sequences showed that there were 19 nucleotide polymorphism loci ( SNPs) and nine parsim-informative sites in the combination sequences. In addition, Ch. indicum showed more obvious sequence polymorphism than those of Ch. indicum (Juhuanao) and Ch. morifolium. The psbA-trnH sequences showed obvious length variation. The NJ Tree showed that Ch. morifolium numbered C2-C5 were clustered into a single subbranch with a bootstrap value of 62%, and Ch. morifolium could be distinguished from Ch. indicum and Ch. indicum (Juhuanao). Moreover, Ch. indicum numbered Z9 and Z10 collected from Gansu province were singly clustered into one branch with a bootstrap value of 77%. It was also found that the changes of psbA-trnH and trnL sequences information of Ch. indicum samples from the northwest were obviously related to the geography and environment. Moreover, Ch.indicum and Ch. indicum (Juhuanao) had obvious differentiation, were also regarded as the evolutionary sources of Ch. morifoliunL Therefore, psbA-trnH+matK+trnL combination sequences as DNA barcode can identify Ch. indicum and Ch. morifolium accurately and rapidly, which provides an important basis for germplasm resources identification and species identification.

关 键 词：野菊 药用菊 DNA条形码 SNPs 鉴定 

分 类 号：R282.5[医药卫生—中药学]