| 摘要: |
| 目的:探讨影响红花SRAP扩增的各种因素,建立能够稳定扩增红花基因组的体系,为研究红花重要性状的遗传基础及建立分子辅助标记育种的技术平台奠定基础.方法:用CTAB法提取红花DNA,设计Taq酶浓度(0.02、0.04、0.06 U/μl)、dNTP浓度(0.15、0.25、0.30 mmol/L)、Primer浓度(0.15、0.30、0.45 μmol/L)3因素3水平27次实验和Mg2+浓度(0.5、1.0、1.5、2.0、2.5、3.0、3.5、4.0 mmol/L)8水平单因素实验.在25 μl体系中加入模板DNA 20 ng.对体系进行优化,用琼脂糖进行检测.结果:本研究建立了适合红花的SRAP体系,Taq酶浓度为0.02 U/μl,dNTP浓度为0.25 mmol/L,Primer 浓度为0.30 μmol/L,Mg2+的浓度为3.0 mmol/L,优化后的体系目标条带增多,重现性好,得到了较好的扩增效果.结论:本研究建立的反应体系适合红花SRAP的研究. |
| 关键词: 红花、相关序列扩增多态性、核酸扩增技术 |
| DOI：10.3724/SP.J.1008.2006.00544 |
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| 基金项目:国家自然科学基金（30271588）. |
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| Establishment and optimization of sequence-related amplified polymorphism amplification system for Carthamus tinctorius L. |
| 彭飒,郭美丽,陈跃华,郭庆华,PENG Sa,GUO Mei-li,CHEN Yue-hua,GUO Qing-hua |
| () |
| Abstract: |
| Objective：To study the factors influencing the amplification of sequence-related amplified polymorphism（SRAP） system in Carthamus tinctorius L. and to establish a stable SRAP reaction system, laying a foundation for molecular marker assistant breeding of Carthamus tinctorius L. Methods： Cetyltrimethylammonium bromide method was used to extract the genomic DNA of Carthamus tinctorius L.. Twenty-seven tests with 3 factors at 3 levels were designed, the conditions including Taq polymerase concentrations （0.02,0.04,0.06 U/μl）, dNTP concentrations （ 0. 15,0. 25,0.30 retool/L） and Primer concentrations （0.15,0.30,0.45 μmol/L） ;another 8 tests were designed based on the single factor of Mg^2＋ concentrations（0.5,1.0,1.5,2.0, 2.5,3.0,3.5 and 4.0 mmol/L）. Twenty ng DNA template was added into 25 μl SRAP reaction system; the system was optimized and agarose electrophoresis was used for determination. Results： A SRAP reaction system for Carthamus tinctorius L. was established. In a 25 μl reaction system, Taq polymerase was 0.02 U/μl, dNTP was 0.25 mmol/L,Primer was 0.30 μmol/ L, and Mg^2＋ was 3.0 mmol/L. Target bands increased after optimization, with good reproducibility, and the amplification results were satisfactory. Conclusion： The SRAP reaction system in this experiment is suitable for analysis of Carthamus tinctorius L. |
| Key words: Carthamus tinctorius L. sequence related amplified polymorphism nucleic acid amplification techniques |
