广东紫珠核糖体DNA ITS序列分子鉴定.docVIP

广东紫珠核糖体DNA ITS序列分子鉴定 　　摘要:采用改良CTAB法提取广东紫珠(Callicarpa kwangtungensis Chun.)植物总DNA,以通用引物ITS4和ITS5 PCR扩增核糖体DNA ITS序列,并运用MEGA5.2软件分析ITS序列和构建系统发育树。结果表明,广东紫珠的核糖体DNA ITS序列长度为658 bp,ITS1和 ITS2的长度分别为229、267 bp,G+C含量分别为67.39%、70.30%。用NJ法根据ITS1与ITS2序列构建系统发育树,ITS片段在种间存在差异,可区分紫珠属中不同的种。测序分析和系统发育树构建,能够对广东紫珠进行准确的分子鉴定,为紫珠属中药材的种类鉴定和种间的分类地位提供分子生物学理论依据。 　　关键词:广东紫珠(Callicarpa kwangtungensis Chun.);ITS序列;分子鉴定 　　中图分类号:Q78文献标识码:A文章编号:0439-8114(2014)10-2435-05 　　 　　 Identification of Callicarpa kwangtungensis Chun. with ITS Sequences 　　of Nuclear Ribosomal DNA 　　 　　LI Jia-min,JIANG Qiong,WANG Jian-ming 　　(College of Chemical and Biological Engineering,Yichun University,Yichun 336000,Jiangxi,China) 　　 　　Abstract: To identify C.kwangtungensis Chun. with ITS of rDNA, the total DNA of the sample was extracted with modified CTAB. The ITS of rDNA was amplified using a pair of primers ITS4 and ITS5 then was cloned, sequenced and analyzed by MEGA5.2. The results showed that the length of ITS of rDNA was 658 bp. The DNA sequence of ITS4 was 229 bp with the content of G+C of 67.39%. The DNA sequence of ITS5 was 267 bp with the content of G+C of 70.30%.The phylogenetic tree based on ITS data was constructed. Molecular identification could be performed on Callicarpa kwangtungensis Chun. It will provid molecular evidence for taxonomy and identification of different species of Callicarpa. 　　Key words: Callicarpa kwangtungensis Chun.;ITS sequences;molecular identification 　　 　　基金项目:江西省教育厅青年基金项目(GJJ10252);江西省天然药物活性成分研究重点实验室开放基金项目 　　 　　广东紫珠(Callicarpa kwangtungensis Chun.)为马鞭草科紫珠亚属多年生落叶植物,适合生长于海拔100~1 000 m的丘陵山地,主要分布于江西、湖南、福建、贵州等省,广东和广西等地也有少量分布。广东紫珠的干燥茎、枝、叶均可入药,具有收敛止血、清热解毒等功效,收载于《中国药典》(2010年版),可用于治疗咯血、便血、肺胃出血、崩漏、肺热咳嗽、咽喉肿痛、烧伤、热毒疮、外伤出血等。化学成分研究表明,广东紫珠富含熊果酸、槲皮素、没食子酸、水杨酸等化学成分[1];药理研究显示具有较强的抑菌作用和显著的止血效果[2],临床上广东紫珠主要用于治疗宫颈糜烂出血、阴道炎、宫颈炎等,是抗宫炎片、抗宫炎颗粒和抗宫炎胶囊等中成药的主要成分[3]。目前,市场上对广东紫珠资源需求量很大,原料药材供不应求,由于大量采挖,野生资源锐减,且遭到严重破坏,加上野生药材质量难以控制,野生资源已不能满足社会需求,对广东紫珠进行资源鉴定和育种成为解决这一问题的关键。准确鉴定广东紫珠有利于指导其育种,因此,有必要在DNA分