| 酚／氯仿抽提法提取绵羊凝血块中基因组DNA |
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| 引用本文： | 曹果清,莫清珊,陈凤仙.酚／氯仿抽提法提取绵羊凝血块中基因组DNA[J].农业科学与技术,2009,10(5):76-78. |
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| 作者姓名： | 曹果清 莫清珊 陈凤仙 |
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| 作者单位： | [1]山西农业大学动物科技学院,山西太谷030801 [2]天津实发冠华生物科技有限公司,天津300462 |
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| 基金项目： | 山西省自然科学基金项目，山西省归国留学人员项目，农业科技成果转化资金项目 |
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| 摘 要： | 目的]对组织DNA提取方法进行改进，建立一种从绵羊凝血块中提取基因组DNA的方法。方法]将绵羊凝血块用眼科剪剪碎，用组织DNA抽提液裂解细胞，用蛋白酶K消化后，经过酚／氯仿抽提，无水乙醇沉淀获得基因组DNA。用NanoDropND-1000微型分光光度计检测DNA浓度和纯度。用0．8％琼脂糖凝胶电泳检验基因组DNA的完整性。以绵羊微卫星位点BM203为扩增位点，分别以F：5’-GG（汀G11：AcAmGTTCCC-3’，R：5’-CTGCTCCCCACTACTCCTTC-3’为上下游引物，进行PCR扩增试验。PCR产物用1．5％琼脂糖凝胶电泳检测．结果]提取的DNA浓度为（159．90±0．70）ng／μl，A260／A280比值为1．80±0．01，分子完整，结果理想。以从凝血块中提取的DNA为模板，对绵羊BM203微卫星位点进行了PCR扩增，扩增产物条带整齐、明亮、特导性强，扩增效果好。结论]该方法简单、实用，提取的DNA可满足后续相关研究对DNA质量的要求，值得推广借鉴。
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| 关 键 词： | 绵羊凝血块 酚／氯仿抽提法 DNA提取 |
Genomic DNA Isolation by Phenol/Chloroform Extracting Method from Sheep Blood Clot |
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| CAO Guo-qing,MO Qing-shan,CHEN Feng-xian.Genomic DNA Isolation by Phenol/Chloroform Extracting Method from Sheep Blood Clot[J].Agricultural Science ＆ Technology,2009,10(5):76-78. |
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| Authors: | CAO Guo-qing MO Qing-shan CHEN Feng-xian |
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| Institution: | 1. College of Animal Science and Veterinary Medicine, Shanxi Agricultural University, Taigu 030801 ; 2. Tianjin Shi-fa Guan-hua Biological Science and Technology Co., Ltd., Tianjin 300462) |
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| Abstract: | Objective] The aim was to establish the method of extracting genomic DNA from sheep blood clot on the basis of the improvement of method for extracting genomic DNA from tissues. Method]The genomic DNA with complete primary structure and high purity was obtained from the sheep blood clot after the steps of cutting the sheep blood clot with ophthalmic scissors, cell lysis with tissue DNA extracts and digested by proteinase K, extracting with phenol/chloroform and precipitating with ethanol were performed. Result] The concentration of the extracted DNA was 159.90 ±0.70 ng/μl and the ratio of the A260/A280 was 1.80 ＋0.01. The sheep microsatellite locus of BM203 was amplified by using the extracted DNA from the sheep blood clot as template of PCR, and the PCR result was perfect. Conclusion]This method is simple and feasible, the quantity and quality of the extracted DNA can satisfy the demands for the subsequent researches. It is worth to extending and using for reference. |
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| Keywords: | Sheep blood clot Phenol/chloroform extracting method DNA extraction |
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