| 一种快速高效制备家蚕微孢子虫基因组DNA和总蛋白的方法 |
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| 引用本文： | 蔡顺风,何欣怡,何祥康,邱海洪,李光才,何永强,鲁兴萌. 一种快速高效制备家蚕微孢子虫基因组DNA和总蛋白的方法[J]. 蚕业科学, 2011, 37(6) |
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| 作者姓名： | 蔡顺风 何欣怡 何祥康 邱海洪 李光才 何永强 鲁兴萌 |
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| 作者单位： | 浙江大学蚕蜂研究所,杭州,310058;浙江出入境检验检疫局技术中心,杭州,310012 |
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| 基金项目： | 现代农业产业技术体系专项(No.CARS-22); 浙江省科技厅重大项目(No.2010C12005-2); 国家质量监督检验检疫总局科研项目(No.2007IK017) |
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| 摘 要： | 家蚕微孢子虫孢子壁坚厚不易破碎,并对许多化学物质有很强的抵抗性,使用常规的核酸和蛋白抽提方法效果不够理想。研究建立玻璃珠破碎法快速制备家蚕微孢子虫基因组DNA和总蛋白的方法,采用琼脂糖凝胶电泳和SDS-PAGE分析所得DNA的完整性和所得总蛋白的差异性,并对提取蛋白进行双向电泳试验。结果表明:使用玻璃珠破碎法对微孢子虫孢子壁的破碎率可达到90%以上,基因组DNA得率约为7.65 fg/粒,总蛋白得率约为788.3 fg/粒,均显著高于常用的发芽法和液氮冻融研磨法的得率。双向电泳结果显示玻璃珠破碎法能有效提取家蚕微孢子虫总蛋白,经考马斯亮蓝染色可检测到约350个蛋白点,蛋白信息丰富,可进一步用于质谱分析。
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| 关 键 词： | 家蚕微孢子虫 核酸 蛋白 提取方法 珠磨式研磨器 双向电泳 |
A Protocol for Fast and Efficient Preparation of Genomic DNA and Total Proteins of Nosema bombycis |
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| CAI Shun-Feng,HE Xin-Yi,HE Xiang-Kang,QIU Hai-Hong,LI Guang-Cai,HE Yong-Qiang,LU Xing-Meng. A Protocol for Fast and Efficient Preparation of Genomic DNA and Total Proteins of Nosema bombycis[J]. Acta Sericologica Sinica, 2011, 37(6) |
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| Authors: | CAI Shun-Feng HE Xin-Yi HE Xiang-Kang QIU Hai-Hong LI Guang-Cai HE Yong-Qiang LU Xing-Meng |
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| Affiliation: | CAI Shun-Feng1 HE Xin-Yi1 HE Xiang-Kang1 QIU Hai-Hong1 LI Guang-Cai1 HE Yong-Qiang2 LU Xing-Meng1(1Institute of Sericulture and Apiculture,Zhejiang University,Hangzhou 310058,China,2Technical Center of Zhejiang Entry-Exit Inspection and Quarantine Bureau,Hangzhou 310012,China) |
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| Abstract: | Because Nosema bombycis has thick rigid wall which is difficult to break and has high endurance to many chemicals,preparation of its DNA and total proteins with conventional methods has no satisfactory effect.Here,we present a protocol for fast and efficient preparation of genomic DNA and total proteins of Nosema bombycis with glass bead beating method.Integrity of DNA and variability of total proteins were assessed by agarose gel electrophoresis and SDS-PAGE analysis.And the extracted proteins were subject... |
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| Keywords: | Nosema bombycis DNA Protein Extraction method BeadBeater 2D-electrophoresis |
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