甜菜CDDP-PCR扩增体系的建立及优化
|
摘要
为了建立最优甜菜的来源保守DNA序列多态(conserved DNA-derived polymorphism,CDDP)分子标记扩增体系,以期利用CDDP分子标记技术进行甜菜品种指纹图谱的构建、核心种质构建及分子标记辅助育种。本实验利用单因素变量的方法对甜菜CDDP体系进行优化;最终确定了甜菜最适CDDP体系,体系总体积为20μL,其中包括10μL MIX,0.5μL引物,0.5μL DNA模板。利用优化的体系对8个甜菜二倍体品系进行扩增,结果表明所有材料都能扩增出清晰稳定的条带,并且多态性丰富,由此可以说明本研究对CDDP进行优化的体系非常适合甜菜CDDP-PCR分析。
The study aims to establish the optimal conserved DNA-derived polymorphism(CDDP)amplification system of sugar beet, with the purpose of using CDDP molecular marker technique to carry outthe sugar beet varieties fingerprint construction, core collection construction and molecular marker assistedbreeding. This experiment optimized the sugar beet CDDP system by using the method of single factor variableand ultimately determined the beet optimum CDDP system with a total volume of 20 μL, containing 10 μL ofMIX, 0.5 μL of primers and 0.5 μL of template DNA. 8 varieties of sugar beets were amplified using theoptimized reaction system and the results showed that all materials could be amplified out clear and stablebands and the primer's polymorphism was very abundant. Thus, the optimized system of CDDP in this study isvery suitable for beet CDDP-PCR analysis.
The study aims to establish the optimal conserved DNA-derived polymorphism(CDDP)amplification system of sugar beet, with the purpose of using CDDP molecular marker technique to carry outthe sugar beet varieties fingerprint construction, core collection construction and molecular marker assistedbreeding. This experiment optimized the sugar beet CDDP system by using the method of single factor variableand ultimately determined the beet optimum CDDP system with a total volume of 20 μL, containing 10 μL ofMIX, 0.5 μL of primers and 0.5 μL of template DNA. 8 varieties of sugar beets were amplified using theoptimized reaction system and the results showed that all materials could be amplified out clear and stablebands and the primer's polymorphism was very abundant. Thus, the optimized system of CDDP in this study isvery suitable for beet CDDP-PCR analysis.
引文
[1]兴旺,潘荣,崔平.甜菜种质资源叶部性状多样性及聚类分析[J].中国农学通报,2015,31(30):155-161.
[2]刘乃新,郭晨晨,孙燕红,等.甜菜SCo T-PCR反应体系的建立及优化[J].中国农学通报,2016,32(34):119-122.
[3]王茂芊,吴则东,王华忠.利用SRAP标记分析中国甜菜三大产区骨干材料的遗传多样性[J].作物学报,2011,37(05):811-819.
[4]吴则东,王茂芊,吴玉梅,等.32个甜菜品种指纹图谱构建与遗传多样性分析[J].中国农学通报,2015,31(12):169-174.
[5]王玉娟,张彦,房经贵,等.利用基于RAPD标记的MCID法快速鉴定72个葡萄品种[J].中国农业科学,2012,45(14):2913-2922.
[6]昝逢刚,应雄美,吴才文,等.98份甘蔗种质资源遗传多样性的AFLP分析[J].中国农业科学,2015,48(05):1002-1010.
[7]段永红,渠云芳,王铭,等.高粱抗丝黑穗病基因SRAP体系优化[J].分子植物育种,2017(02):611-617.
[8]王瑞云,季煦,陆平,等.利用荧光SSR分析中国糜子遗传多样性[J].作物学报,2017(04):530-548.
[9]李玉阁,苏亚中,张大乐,等.中国节节麦基于ISSR标记的遗传多样性分析[J].麦类作物学报,2017,37(01):30-39.
[10]宁洽,刘文国,杨伟光,等.SNP标记在玉米研究上的应用进展[J].玉米科学,2017,25(01):57-61.
[11]胡建斌,张帆,李贞煌,等.黄瓜DAMD反应体系的建立及种质遗传资源研究[J].西北植物学报,2010,30(04):652-658.
[12]李强,苏二虎,高聚林,等.大豆CDDP分子标记技术体系的优化、验证及引物筛选[J].大豆科学,2013,32(03):310-315.
[13]李田,郭俊娥,郑成淑,等.菊花品种的遗传多样性分析及CDDP指纹图谱构建[J].北京林业大学学报,2014,36(04):94-101.
[14]李莹莹.牡丹CDDP分子标记引物筛选及多态性分析[J].核农学报,2013,27(08):1099-1105.
[15]房文秀,夏秀英,安利佳,等.越橘种质资源的CDDP遗传多样性及聚类分析[J].中国农学通报,2016,32(28):136-143.
[16]Collard B C Y,Mackill D J.Conserved DNA-Derived Polymorphism(CDDP):A Simple and Novel Method for Generating DNA Markers in Plants[J]..Plant Molecular Biology Reporter.2009,27(4):558-562.
[17]Talebi R.Comparative Efficiency of Functional Gene-based Markers,Start Codon Targeted Polymorphism(SCo T)and Conserved DNA-derived Polymorphism(CDDP)with ISSR Markers for Diagnostic Fingerprinting in Wheat(Triticum aestivum L.)[J].Cereal Research Communications.2014(4):365-373.
[18]Hajibarat Z,Saidi A,Hajibarat Z,et al.Characterization of genetic diversity in chickpea using SSR markers,Start Codon Targeted Polymorphism(SCo T)and Conserved DNA-Derived Polymorphism(CDDP)[J].Physiology and Molecular Biology of Plants,2015,21(3):1-9.
[19]熊发前,唐荣华,庄伟建,等.一种基于表达序列标签(EST)的新型目标分子标记技术—保守区域扩增多态性(Co RAP)[J].广西农业科学,2010,02:100-103.
[20]熊发前,唐荣华,陈忠良,等.目标起始密码子多态性(SCo T):一种基于翻译起始位点的目的基因标记新技术[J].分子植物育种,2009,03:635-638.
[21]Poczai P,Varga I,Bell N E,et al.Genetic diversity assessment of bittersweet(Solanum dulcamara,Solanaceae)germplasm using conserved DNA-derived polymorphism and intron-targeting markers[J].Annals of Applied Biology,2011,159:141-153.
[22]Poczai P,Varga I,Laos M,et al.Advances in plant gene-targeted and functional markers:a review[J].Plant Methods,2013,9:6.
[23]胡根海,喻树迅.利用改良的CTAB法提取棉花叶片总RNA[J].棉花学报,2007,19(01):69-70.
[24]罗霆,杨海霞,岑华飞,等.SCo T分子标记在割手密遗传图谱构建中的应用[J].植物遗传资源学报,2013,14(4):704-710.
[25]熊发前,蒋菁,钟瑞春,等.两种新型目标分子标记技术—CDDP与PAAP[J].植物生理学通讯,2010(09):871-875.
[2]刘乃新,郭晨晨,孙燕红,等.甜菜SCo T-PCR反应体系的建立及优化[J].中国农学通报,2016,32(34):119-122.
[3]王茂芊,吴则东,王华忠.利用SRAP标记分析中国甜菜三大产区骨干材料的遗传多样性[J].作物学报,2011,37(05):811-819.
[4]吴则东,王茂芊,吴玉梅,等.32个甜菜品种指纹图谱构建与遗传多样性分析[J].中国农学通报,2015,31(12):169-174.
[5]王玉娟,张彦,房经贵,等.利用基于RAPD标记的MCID法快速鉴定72个葡萄品种[J].中国农业科学,2012,45(14):2913-2922.
[6]昝逢刚,应雄美,吴才文,等.98份甘蔗种质资源遗传多样性的AFLP分析[J].中国农业科学,2015,48(05):1002-1010.
[7]段永红,渠云芳,王铭,等.高粱抗丝黑穗病基因SRAP体系优化[J].分子植物育种,2017(02):611-617.
[8]王瑞云,季煦,陆平,等.利用荧光SSR分析中国糜子遗传多样性[J].作物学报,2017(04):530-548.
[9]李玉阁,苏亚中,张大乐,等.中国节节麦基于ISSR标记的遗传多样性分析[J].麦类作物学报,2017,37(01):30-39.
[10]宁洽,刘文国,杨伟光,等.SNP标记在玉米研究上的应用进展[J].玉米科学,2017,25(01):57-61.
[11]胡建斌,张帆,李贞煌,等.黄瓜DAMD反应体系的建立及种质遗传资源研究[J].西北植物学报,2010,30(04):652-658.
[12]李强,苏二虎,高聚林,等.大豆CDDP分子标记技术体系的优化、验证及引物筛选[J].大豆科学,2013,32(03):310-315.
[13]李田,郭俊娥,郑成淑,等.菊花品种的遗传多样性分析及CDDP指纹图谱构建[J].北京林业大学学报,2014,36(04):94-101.
[14]李莹莹.牡丹CDDP分子标记引物筛选及多态性分析[J].核农学报,2013,27(08):1099-1105.
[15]房文秀,夏秀英,安利佳,等.越橘种质资源的CDDP遗传多样性及聚类分析[J].中国农学通报,2016,32(28):136-143.
[16]Collard B C Y,Mackill D J.Conserved DNA-Derived Polymorphism(CDDP):A Simple and Novel Method for Generating DNA Markers in Plants[J]..Plant Molecular Biology Reporter.2009,27(4):558-562.
[17]Talebi R.Comparative Efficiency of Functional Gene-based Markers,Start Codon Targeted Polymorphism(SCo T)and Conserved DNA-derived Polymorphism(CDDP)with ISSR Markers for Diagnostic Fingerprinting in Wheat(Triticum aestivum L.)[J].Cereal Research Communications.2014(4):365-373.
[18]Hajibarat Z,Saidi A,Hajibarat Z,et al.Characterization of genetic diversity in chickpea using SSR markers,Start Codon Targeted Polymorphism(SCo T)and Conserved DNA-Derived Polymorphism(CDDP)[J].Physiology and Molecular Biology of Plants,2015,21(3):1-9.
[19]熊发前,唐荣华,庄伟建,等.一种基于表达序列标签(EST)的新型目标分子标记技术—保守区域扩增多态性(Co RAP)[J].广西农业科学,2010,02:100-103.
[20]熊发前,唐荣华,陈忠良,等.目标起始密码子多态性(SCo T):一种基于翻译起始位点的目的基因标记新技术[J].分子植物育种,2009,03:635-638.
[21]Poczai P,Varga I,Bell N E,et al.Genetic diversity assessment of bittersweet(Solanum dulcamara,Solanaceae)germplasm using conserved DNA-derived polymorphism and intron-targeting markers[J].Annals of Applied Biology,2011,159:141-153.
[22]Poczai P,Varga I,Laos M,et al.Advances in plant gene-targeted and functional markers:a review[J].Plant Methods,2013,9:6.
[23]胡根海,喻树迅.利用改良的CTAB法提取棉花叶片总RNA[J].棉花学报,2007,19(01):69-70.
[24]罗霆,杨海霞,岑华飞,等.SCo T分子标记在割手密遗传图谱构建中的应用[J].植物遗传资源学报,2013,14(4):704-710.
[25]熊发前,蒋菁,钟瑞春,等.两种新型目标分子标记技术—CDDP与PAAP[J].植物生理学通讯,2010(09):871-875.