[1]孙祖霞,刘兆磊*,陈素梅,等.荷花SRAP-PCR反应体系的优化与确立[J].南京农业大学学报,2011,34(6):53.[doi:10.7685/j.issn.1000-2030.2011.06.010]
 SUN Zu-xia,LIU Zhao-lei*,CHEN Su-mei,et al.Optimization and establishment of SRAP-PCR reaction system for Nelumbo nucifera Gaertn[J].Journal of Nanjing Agricultural University,2011,34(6):53.[doi:10.7685/j.issn.1000-2030.2011.06.010]
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荷花SRAP-PCR反应体系的优化与确立()
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《南京农业大学学报》[ISSN:1000-2030/CN:32-1148/S]

卷:
34卷
期数:
2011年6期
页码:
53
栏目:
OA栏目
出版日期:
2011-11-30

文章信息/Info

Title:
Optimization and establishment of SRAP-PCR reaction system for Nelumbo nucifera Gaertn
作者:
孙祖霞1刘兆磊1*陈素梅1陈发棣1楼望淮1郭海林12
1.南京农业大学园艺学院,江苏 南京 210095;2.江苏省中国科学院植物研究所,江苏 南京 210014
Author(s):
SUN Zu-xia1LIU Zhao-lei1*CHEN Su-mei1CHEN Fa-di1LOU Wang-huai1GUO Hai-lin12
1.College of Horticulture,Nanjing Agricultural University,Nanjing 210095,China;2.Institute of Botany,Jiangsu Province and Chinese Academy of Sciences,Nanjing 210014,China
关键词:
荷花SRAP-PCR正交试验设计反应体系优化
Keywords:
Nelumbo nucifera GaertnSRAP-PCRorthogonal experiment designoptimization of reaction system
分类号:
S682.32
DOI:
10.7685/j.issn.1000-2030.2011.06.010
摘要:
以荷花(Nelumbo nucifera Gaertn)品种‘新红’叶片为材料,采用L16(45)正交试验设计,对SRAP-PCR反应体系中的Mg2+、dNTPs、TaqDNA聚合酶、引物和模板DNA用量5因素进行了优化,并确立了适用于荷花SRAP-PCR的最佳反应体系。结果表明:荷花的SRAP-PCR最佳反应体系为:反应总体积10 μL,包含2.0 mmol·L-1 Mg2+、300 μmol·L-1 dNTPs、0.5 U TaqDNA聚合酶、4 μmol·L-1上下游引物、50 ng DNA及10×PCR Buffer。各因素水平变化对反应体系影响由大到小依次为:TaqDNA聚合酶、Mg2+、引物、dNTPs、DNA。用48个荷花品种对优化的SRAP-PCR反应体系进行验证,均获得了条带清晰、多态性丰富的扩增图谱,证实了该体系的稳定性和适应性。
Abstract:
Using Nelumbo nucifera Gaertn ‘Xinhong’ leaves as the experimental material,by the orthogonal experiment design L16(45),five factors including concentration for Mg2+ and dNTPs,dosage for TaqDNA polymerase,primer and template DNA in SRAP-PCR reaction system were optimized,and the optimization SRAP-PCR reaction system suitable for lotus were also established.The result showed that the optimal SRAP-PCR reaction system was as follows:total volume 10 μL,including 2.0 mmol·L-1 Mg2+,300 μmol·L-1 dNTPs,0.5 U TaqDNA polymerase,4 μmol·L-1 primer,50 ng DNA and 10×PCR Buffer.The order of each factor in different levels affecting the result of PCR was:TaqDNA polymerase,Mg2+,primer,dNTPs,DNA.The optimal SRAP-PCR reaction system was identified by means of genomic DNA from 48 varieties of N.nucifera Gaertn and the amplification pattern with clear band and rich polymorphism was obtained.It is concluded that the SRAP-PCR reaction system is steady and reliable.

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备注/Memo

备注/Memo:
收稿日期:2011-06-09基金项目:国家公益性行业(农业)科研专项(200903020)作者简介:孙祖霞,硕士研究生。*通讯作者,刘兆磊,副教授,硕导,主要从事观赏植物种质资源与生物技术研究,E-mail:lzl@njau.edu.cn。
更新日期/Last Update: 2011-12-06