绞股蓝的RAPD指纹图谱鉴定及其特异DNA片段克隆、序列分析  

Fingerprints Identification of Gynostemma pentaphyllum by RAPD and Cloning and Analysis of Its Specific DNA Fragment

作　　者：蒋军富[1] 李雄英[1] 吴耀生[1] 罗育[1] 赵瑞强[1] 蓝秀万[1] 

机构地区：[1]广西医科大学生物化学与分子生物学教研室,广西南宁530021

出　　处：《中药材》2009年第2期190-193,共4页Journal of Chinese Medicinal Materials

基　　金：广西科学基金(桂科基0575065;桂科基0731065);广西区教育厅硕士研究生科研创新项目(2006105981001M02);广西大型仪器协作共用网大型仪器测试补助(4841-2007-063)

摘　　要：目的：在DNA分子水平上鉴定绞股蓝及其伪品乌蔹莓。方法：用筛选出的两条随机引物（WGS001，WGS004）对不同采集地绞股蓝及乌蔹莓基因组DNA进行PCR扩增，将引物WGS004扩增7份绞股蓝样品中的具有相同迁移率的1条500bp左右的DNA片段进行克隆、测序、一致性分析，并在NCBI上作Blastn比对检索。结果：以上两条引物能分别扩增得到绞股蓝与乌蔹莓样品基因组的差异性DNA片段，这些片段可以明确区别绞股蓝与乌蔹莓。经克隆获得的7条绞股蓝DNA序列问的一致性在45．7％～94．5％之间，在NCBI上检索后未见相似序列的报道，为新序列。结论：RAPD技术可有效地鉴别绞股蓝和乌蔹莓，本研究克隆得到一段绞股蓝DNA序列，为绞股蓝的品种鉴定、辅助选择育种等研究奠定了基础。Objective ： To identify the resources of Gynostemma pentaphyllum and its spurious breed plant Cayratia japonica at level of DNA. Methods： Two random primers（WGS001 ,WGS004）screened were applied to do random amplification with genomic DNA extracted from Gynostemma pentaphyllum and Cayratia japonica which were collected from different habitats. After amplificated with WGS004, one characteristic fragment about 500 bp which was common to all Gynostemma pentaphyllum samples studied but not to Cayratia japonica was cloned and sequenced. Then these sequences obtained were analyzed for identity and compared by Blastn program in GenBank. Results： There were obvious different bands amplified by above two primers in their fingerprints of genomic DNA. On the basis of these different bands of DNA fingerprints, they could distinguish Gynostemma pentaphyllum and Cayratia japonica obviously. Sequence alignment of seven cloned bands showed that their identities ranged from 45.7%～94. 5%. There was no similar genome sequences searched in GenBank. This indicated that these seven DNA fragments had not been reported before and they should be new sequences. Conclusion ： RAPD technique can be used for the accurate identification of Gynostemma pentaphyllum and its counterfeit goods Cayratia japonica. Besides, these specific DNA sequences for Gynostemrna pentaphyllum in this study are useful for the further research on identification of species and assisted selection breeding in Gynostemma pentaphyllum.

关 键 词：绞股蓝 RAPD 鉴别 特异DNA片段 克隆 

分 类 号：R282.5[医药卫生—中药学]