| 丝状真菌顶头孢霉染色体DNA的提取 |
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| 引用本文： | 徐威,朱春宝,朱宝泉,姚新生. 丝状真菌顶头孢霉染色体DNA的提取[J]. 沈阳药科大学学报, 2004, 21(3): 226-229,236 |
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| 作者姓名： | 徐威 朱春宝 朱宝泉 姚新生 |
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| 作者单位： | 1. 沈阳药科大学制药工程学院,辽宁,沈阳,110016
2. 上海医药工业研究院,上海,200040 |
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| 基金项目： | 国家高技术研究发展计划 (2 0 0 1AA2 14 2 0 1) |
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| 摘 要： | 目的探索提取头孢菌素C产生菌顶头孢霉基因组DNA的方法。方法采用氯化苄法、液氮研磨法和酶裂解法。结果较其他两种方法比 ,氯化苄法提取效果好。当提取液pH值为 9 0～10 0、EDTA浓度为 12 0mmol·L-1时 ,所得的顶头孢霉染色体DNA的质量和数量均较理想 ,每克(湿重 )顶头孢霉菌丝体能得到 96 μg的染色体DNA。常规琼脂糖凝胶电泳分析 ,其DNA片断大于2 3kb。结论该分离方法获得的染色体DNA质量较高 ,可进行限制性内切酶酶解并适合后续特定基因的PCR扩增反应
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| 关 键 词： | 顶头孢霉菌 基因组DNA 提取 研磨 细胞壁 多聚酶链式反应 |
| 文章编号： | 1006-2858(2004)03-0226-04 |
A new method for isolation of chromosomal DNA from filamentous fungus Cephalosporium acremonium |
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| XU Wei ,ZHU Chun-bao ,ZHU Bao-quan ,YAO Xin-sheng. A new method for isolation of chromosomal DNA from filamentous fungus Cephalosporium acremonium[J]. Journal of Shenyang Pharmaceutical University, 2004, 21(3): 226-229,236 |
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| Authors: | XU Wei ZHU Chun-bao ZHU Bao-quan YAO Xin-sheng |
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| Affiliation: | XU Wei 1,ZHU Chun-bao 2,ZHU Bao-quan 2,YAO Xin-sheng 1 |
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| Abstract: | Objective To investigate the genomic DNA extraction methods of Cephalosporium acremonium which produce Cephalosporin C.Methods Three methods including benzyl chloride,Homogenizing in liquid nitrogen and enzymolysis were used.Results The results showed that DNA extraction method with benzyl chloride was better than the other two methods.The amount of chromosomal DNA isolated could reach as high as 96 μg per gram wet mycelia when the pH and the concentration of EDTA in the extraction buffer were kept between 9.0～10.0 and 120 mmol·L -1 and its molecular weight is over 23 kb by agarose gel electrophoresis.Conclusion The isolated DNA is good enough for enzyme digestion and the following PCR amplification. |
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| Keywords: | Cephalosporium acremonium genomic DNA extraction Homogenizing cell wall PCR |
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