适于草果遗传多样性分析的RAPD引物筛选
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摘要
以云南省文山州西畴县和德宏州陇川县的12株草果优良无性系为材料,利用RAPD标记技术,选用192个RAPD随机引物,对草果进行PCR扩增,并用UPGMA法对RAPD引物扩增条带进行聚类分析,计算其遗传相似系数,以期筛选出适用于草果遗传多样性分析的RAPD有效引物。实验共筛选出21个条带清晰、多态性丰富并具有可重复性特点的有效引物。21个引物在实验选用的12份样品中共扩增出177条DNA带,其中多态性带数为119条,占扩增总带数的67.2%,每个引物平均扩增出8.4条带。结果表明,12份草果样品聚为A组和B组,其中A组又分为两个亚组,聚类结果与所选用材料的地理来源基本一致,这也说明该研究所筛选得到的21个RAPD引物适用于草果的遗传多样性分析。该研究为应用RAPD标记技术对草果进行遗传多样性分析等研究奠定了基础。
In this study,192 RAPD random primers used to amplify the PCR of Amomum tsao-ko,and the aim was to screen out RAPD effective primers suitable for genetic diversity analysis of A. tsao-ko. In the end,21 effective primers with clear bands,rich polymorphism and good reproducibility were screened out. And 21 primers were amplified 177 DNA bands in 12 samples,of which the number of polymorphic bands was 119,accounting for67. 2% of the total bands,and an average of 8. 4 ribbons for each primer. UPGMA method used to cluster analysis,and then calculated the genetic similarity coefficient about A. tsao-ko. The results showed that 12 samples of A. tsao-ko were divided into group A and group B,and group A was divided into two subgroups. The results of cluster analysis were consistent with the geographical sources of the selected materials,which indicated that the 21 RAPD primers screened in this study suitable for the genetic diversity analysis of A. tsao-ko. This research has laid a foundation for the future researchers to use RAPD marker technology to analyze the genetic diversity of A. tsao-ko.
In this study,192 RAPD random primers used to amplify the PCR of Amomum tsao-ko,and the aim was to screen out RAPD effective primers suitable for genetic diversity analysis of A. tsao-ko. In the end,21 effective primers with clear bands,rich polymorphism and good reproducibility were screened out. And 21 primers were amplified 177 DNA bands in 12 samples,of which the number of polymorphic bands was 119,accounting for67. 2% of the total bands,and an average of 8. 4 ribbons for each primer. UPGMA method used to cluster analysis,and then calculated the genetic similarity coefficient about A. tsao-ko. The results showed that 12 samples of A. tsao-ko were divided into group A and group B,and group A was divided into two subgroups. The results of cluster analysis were consistent with the geographical sources of the selected materials,which indicated that the 21 RAPD primers screened in this study suitable for the genetic diversity analysis of A. tsao-ko. This research has laid a foundation for the future researchers to use RAPD marker technology to analyze the genetic diversity of A. tsao-ko.
引文
[1]国家药典委员会.中华人民共和国药典(二部)[M].北京:中国医药科技出版社,2010:837-841.
[2]中国科学院昆明植物研究所.云南植物志[M].北京:科学出版社,1997.
[3]Yang Y,Yue Y,Runwei Y,et al.Cytotoxic,apoptotic and antioxidant activity of the essential oil of Amomum tsao-ko[J].Bioresour Technol,2010,101(11):4205-4211.
[4]吴燕飞,葛发欢,史庆龙,等.超临界CO2萃取草果挥发油成分研究[J].中药材,1997,20(5):240-241.
[5]谢小梅,龙凯,钟裔荣,等.高良姜、草果防霉作用的实验研究[J].中国药业,2002,11(5):45-46.
[6]张薇,杨生超,魏翔,等.云南草果种植发展现状及对策[J].世界科学技术(中医药现代化),2011,13(5):899-903.
[7]严美荣.草果遗传多样性分析及其混淆品的ITS序列差异比较研究[D].昆明:云南中医学院,2012.
[8]陈云鹏,曹家树,缪颖,等.芸薹类蔬菜基因组DNA遗传多样性的RAPD分析[J].浙江大学学报(农业与生命科学版),2000,26(2):131-136.
[9]方宣钧,黄育民,陈启锋,等.若干水稻品种(组合)的等位酶和RAPD遗传分析[J].中国农业科学,1999,32(2):1-8.
[10]许东河,高忠,盖均镒,等.中国野生大豆与栽培大豆等位酶、RFLP和RAPD标记的遗传多样性与演化趋势分析[J].中国农业科学,1999,32(6):16-22.
[11]Charles J S,Fred J M.A quick and versatile method for plant DNA extraction from small amounts of tissue[J].Nucleic Acids Research,1990,18:6531-6535.
[12]张忠廉,李学兰,杨春勇,等.砂仁遗传多样性的ISSR分析[J].中草药,2011,42(3):570-574.
[13]陶正明,冷春鸿,吴志刚,等.温郁金遗传多样性的ISSR分析[J].浙江农业学报,2009,21(3):207-210.
[14]田蜜.川产姜黄的种质资源评价及优良株系初步筛选研究[D].成都:成都中医药大学,2009.
[15]高德民,刘振伟,樊守金.姜品种遗传多样性的RAPD分析[J].农业生物技术学报,2006(2):245-249.
[16]徐国钧,徐珞珊,王峥涛,等.常用中药材品种整理和研究(第-2)[M].福州:福建科学技术出版社,1997.
[17]唐德英,马洁,里二,等.我国草果栽培技术研究概况[J].亚太传统医药,2009,5(7):157-162.
[18]罗晓霞.草果与拟草果的性状与显微鉴别[J].中药材,2014,37(2):227-229.
[19]余南才,叶明波.草果与草豆蔻的鉴别比较[J].中国医院药学杂志,1994(4):183-184.
[20]高学昌.草果及其伪品印度砂仁的鉴别[J].中药通报,1986(4):24-25.
[2]中国科学院昆明植物研究所.云南植物志[M].北京:科学出版社,1997.
[3]Yang Y,Yue Y,Runwei Y,et al.Cytotoxic,apoptotic and antioxidant activity of the essential oil of Amomum tsao-ko[J].Bioresour Technol,2010,101(11):4205-4211.
[4]吴燕飞,葛发欢,史庆龙,等.超临界CO2萃取草果挥发油成分研究[J].中药材,1997,20(5):240-241.
[5]谢小梅,龙凯,钟裔荣,等.高良姜、草果防霉作用的实验研究[J].中国药业,2002,11(5):45-46.
[6]张薇,杨生超,魏翔,等.云南草果种植发展现状及对策[J].世界科学技术(中医药现代化),2011,13(5):899-903.
[7]严美荣.草果遗传多样性分析及其混淆品的ITS序列差异比较研究[D].昆明:云南中医学院,2012.
[8]陈云鹏,曹家树,缪颖,等.芸薹类蔬菜基因组DNA遗传多样性的RAPD分析[J].浙江大学学报(农业与生命科学版),2000,26(2):131-136.
[9]方宣钧,黄育民,陈启锋,等.若干水稻品种(组合)的等位酶和RAPD遗传分析[J].中国农业科学,1999,32(2):1-8.
[10]许东河,高忠,盖均镒,等.中国野生大豆与栽培大豆等位酶、RFLP和RAPD标记的遗传多样性与演化趋势分析[J].中国农业科学,1999,32(6):16-22.
[11]Charles J S,Fred J M.A quick and versatile method for plant DNA extraction from small amounts of tissue[J].Nucleic Acids Research,1990,18:6531-6535.
[12]张忠廉,李学兰,杨春勇,等.砂仁遗传多样性的ISSR分析[J].中草药,2011,42(3):570-574.
[13]陶正明,冷春鸿,吴志刚,等.温郁金遗传多样性的ISSR分析[J].浙江农业学报,2009,21(3):207-210.
[14]田蜜.川产姜黄的种质资源评价及优良株系初步筛选研究[D].成都:成都中医药大学,2009.
[15]高德民,刘振伟,樊守金.姜品种遗传多样性的RAPD分析[J].农业生物技术学报,2006(2):245-249.
[16]徐国钧,徐珞珊,王峥涛,等.常用中药材品种整理和研究(第-2)[M].福州:福建科学技术出版社,1997.
[17]唐德英,马洁,里二,等.我国草果栽培技术研究概况[J].亚太传统医药,2009,5(7):157-162.
[18]罗晓霞.草果与拟草果的性状与显微鉴别[J].中药材,2014,37(2):227-229.
[19]余南才,叶明波.草果与草豆蔻的鉴别比较[J].中国医院药学杂志,1994(4):183-184.
[20]高学昌.草果及其伪品印度砂仁的鉴别[J].中药通报,1986(4):24-25.