| 狗牙根SRAP-PCR反应体系的优化 |
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| 引用本文： | 黄春琼,;周少云,;刘国道,;白昌军.狗牙根SRAP-PCR反应体系的优化[J].广西农业生物科学,2009(5):981-989. |
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| 作者姓名： | 黄春琼 ;周少云 ;刘国道 ;白昌军 |
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| 作者单位： | [1]中国热带农业科学院热带作物品种资源研究所,儋州571737; [2]农业部热带种质资源利用开放重点实验室,儋州571737; [3]海南大学农学院,儋州571737 |
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| 基金项目： | 基金项目：本研究由农业部973项（2007CB108903）、国家科技支撑计划项目（2008BADB3B08）和热带牧草种质资源保护项目共同资助 |
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| 摘 要： | 本研究以狗牙根幼叶为为试材,采用单因子试验和正交设计试验2种方法,对SRAP-PCR反应体系进行优化试验,结果表明狗牙根25μLSRAP-PCR最佳反应体系为：1.5mmol/LMg2＋、0.25mmol/LdNTP、0.4μmol/L引物、1.5UTaq酶和80ng模板DNA,最佳退火温度为50.6℃。运用该体系对30份狗牙根种质进行验证,电泳结果显示扩增条带清晰、多态性高,证明该体系稳定可靠,这一优化体系的建立为今后利用SRAP标记技术对狗牙根进行分子生物学研究提供了帮助。
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| 关 键 词： | 狗牙根 SRAP-PCR 单因子试验 正交设计 体系优化 |
The Optimization of SRAP-PCR System of Bermuda Grass |
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| Institution: | Huang Chunqiong Zhou Shaoyun Liu Guodao Bai Changjun (1 Tropical Crop Genetic Resources Institute, CATAS, Danzhou, 571737; 2 Key Laboratory for Tropical Crop Germplasm Utilization, Ministry of Agriculture, Danzhou, 571737; 3 College of Agriculture, Hainan University, Danzhou, 571737) |
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| Abstract: | In this paper, we attempted to establish the optimal SRAP-PCR amplification system of bermuda grass by employing single factor tests and orthogonal design approaches and using young leaves as tested material. According to our experiment, we established a SRAP-PCR system most suitable for bermuda grass, which came out total 25 μL reaction system containing 1.5 mmol/L Mg^2＋, 0.25 mmol/L dNTP, 0.4 μmol/L primer, 1.5 U Taq DNA polymerase, and 80 ng template DNA. The optimum annealing temperature was 50.6℃. We validated 30 bermuda grass germplasms by using this SRAP-PCR system, and the electrophoresis results showed that the amplification bands were clear and highly polymorphic, which proved that this system was stable and reliable. This optimized SRAP-PCR system might facilitate to be as one of the marker based PCR protocols applying for further molecular genetics research on bermuda grass. |
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| Keywords: | Bermuda grass SRAP-PCR Single factor test Orthogonal design Optimized system |
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