褐黄孢链霉菌基因组DNA的提取及RAPD体系的优化

刘丹,骆健美,刘峰,王敏

中国生物工程杂志 ›› 2008, Vol. 28 ›› Issue (专刊) : 172-177.

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PDF(527 KB)
中国生物工程杂志 ›› 2008, Vol. 28 ›› Issue (专刊) : 172-177.
论文

褐黄孢链霉菌基因组DNA的提取及RAPD体系的优化

  • 刘丹 骆健美 刘峰 王敏
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Genomic DNA extraction and optimization of RAPD amplifying conditions of Streptomyces gilvosporeus

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摘要

以改进的SDS法提取褐黄孢链霉菌基因组DNA,对其进行随机扩增多态性(RAPD)研究。通过单因素和正交试验相结合的方法,建立了适于褐黄孢链霉菌RAPD分析的PCR反应体系:包括Taq聚合酶、Mg2+、随机引物、dNTPs和模板浓度。在20μL体系中,模板DNA浓度60-150ng,Taq聚合酶为1.0-1.5U,引物浓度0.3-0.4mmol/L,dNTPs浓度200-250μmol/L,Mg2 +浓度2.5-3.0mmol/L。在此反应体系下可扩增出条带数目多且清晰稳定的电泳图谱。

Abstract

An improved SDS method was used to extract genomic DNA from Streptomyces gilvosporeus. The RAPD conditions were optimized by single factor experiment and orthogonal experiment, including Taq DNA polymerase, Mg2+, primer , dNTPs and template DNA. The results were as follows: DNA 60-150 ng, Taq DNA polymerase 1.0-1.5U, primer concentration 0.3-0.4 mmol/L, dNTPs concentration 200-250 μmol/L, Mg2+ concentration 2.5-3.0 mmol/L in 20μL PCR system. Under above optimal conditions,the abundant, stable and clear strips could be obtained.

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刘丹,骆健美,刘峰,王敏. 褐黄孢链霉菌基因组DNA的提取及RAPD体系的优化[J]. 中国生物工程杂志, 2008, 28(专刊): 172-177
Genomic DNA extraction and optimization of RAPD amplifying conditions of Streptomyces gilvosporeus[J]. China Biotechnology, 2008, 28(专刊): 172-177

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