| 青钱柳ISSR-PCR反应体系的优化 |
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| 引用本文： | 谈探,金则新,李钧敏.青钱柳ISSR-PCR反应体系的优化[J].安徽农业科学,2006,34(24):6450-6451. |
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| 作者姓名： | 谈探 金则新 李钧敏 |
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| 作者单位： | 北京林业大学自然保护区学院,北京,100083;台州学院生态研究所,浙江临海,317000;台州学院生态研究所,浙江临海,317000 |
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| 基金项目： | 浙江省自然科学基金资助项目(Y504220) |
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| 摘 要： | 采用改进的SDS法提取青钱柳基因组DNA,测试青钱柳ISSR扩增的最适退火温度,并采用单因素试验,测试了模板DNA、Mg2+、dNTP、BSA、引物\Taq酶6个因素对青钱柳ISSR扩增的影响。结果表明:合适的退火温度为56.3℃;适宜的扩增体系为:10μlPCR反应体积中,1×Taq酶配套缓冲液浓度为(10 mmol/LTris.HCl,pH值9.0,浓度为50 mmol/LKCl,0.1%Triton X-100),12 ng模板DNA,浓度为1.5 mmol/LMgCl2,浓度为1.0 mmol/L4×dNTP,1mg/ml BSA,15 pmol引物,0.75 UTaq酶。
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| 关 键 词： | 青钱柳 ISSR 反应体系 优化 |
| 文章编号： | 0517-6611（2006）24-6450-02 |
| 收稿时间： | 09 17 2006 12:00AM |
| 修稿时间： | 2006-09-17 |
Optimization of the ISSR-PCR Reaction System of Cyclocarya paliurus |
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| TAN Tan et al.Optimization of the ISSR-PCR Reaction System of Cyclocarya paliurus[J].Journal of Anhui Agricultural Sciences,2006,34(24):6450-6451. |
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| Authors: | TAN Tan |
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| Abstract: | The genomic DNA of Cyclocarya paliurus was extracted with the improved SDS method.Bsed on its DNA,the ISSR amplificatory reaction system was optimized.The fitting annealing temperature was confirmed and the effect of Mg2+ concentration,dNTP concentration,BSA dosage,primer dosage,Taq DNA polymerase dosage and DNA templates dosage on ISSR amplification were tested with the single factor method.The result was that,the fitting annealing temperature was 56.3 ℃ and the optimal reaction system of ISSR for Cyclocarya paliurus was determined as follows: 1×Taq polymerase corresponding buffer(10 mmol/LTris·HCl pH 9.0,50 mmol/L KCl,0.1 % Triton X-100),12ng template DNA,1.5mmol/L MgCl2,1.0 mmol/L 4×dNTP,1mg/ml BSA,15 pmol primer,0.75U Taq DNA polymerase in total 10 μl reaction volume.Thus,the establishment of ISSR-PCR reaction system settled the genetic diversity foundation for Cyclocarya paliurus. |
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| Keywords: | Cyclocarya paliurus ISSR Reaction system Optimization |
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