摘要
从培养时间、裂解溶液、抽提和沉淀时间等方面对苏云金芽胞杆菌(Bacillus thuringiensis,Bt)基因组DNA提取技术进行改进。提取8株对蛴螬有杀虫活性的野生Bt菌株的基因组DNA,分析其纯度,并进行PCR分析与酶切分析。试验结果表明,新的提取方法耗时36 min,明显短于旧方法所用时间(97 min);两种方法提取的基因组DNA A260/A280均大于1.8,无明显区别;琼脂糖凝胶电泳结果显示,上样量相同时,新方法提取的基因组DNA浓度是旧方法的5倍以上;新方法提取的基因组DNA能作为模板灵敏地扩增出测试基因,所提取的DNA能被限制性内切酶完全酶切,证明DNA具有很高的纯度。本研究改进的基因组DNA提取方法耗时短、产量高,并能满足PCR扩增、酶切等分子生物学需要。
Extraction technique of genomic DNA form Bacillus thuringiensis was improved with regard to culturing time,lysis solution,extraction and precipitation time in this study.Genomic DNA from eight Bacillus thuringiensis isolates toxic to scarab larvae were obtained by this modified method,and purity of these DNA samples was assayed,then PCR amplification,restriction endonuclease enzyme reaction were conducted respectively.The result indicated that 36 min was needed for the new method,while 97 min needed for the old method;The A260/A280 value of the both DNA samples was beyond 1.8,no difference between them.Agarose gel electrophoresis analysis result showed DNA output of the new method was more than five times higher than that of the old one;The test gene could be amplified easily from DNA sample extracted by the new method,and restriction reaction result indicated the genomic DNA obtained by modified method was digested completely,demonstrating high purity of this DNA sample.The modified method is prominent on short time,high yield and purity,the DNA sample obtained can be used for many molecular biology studies,including PCR amplification and restriction endonuclease digestion.
出处
《生物技术通报》
CAS
CSCD
北大核心
2012年第11期197-201,共5页
Biotechnology Bulletin
基金
国家自然科学基金项目(31171911)
国家"973"计划项目(2009CB118902)
国家"863"计划项目(2011AA10A203)
关键词
苏云金芽胞杆菌
基因组DNA
提取
纯度
PCR
酶切
Bacillus thuringiensis Genomic DNA Extraction Purification PCR Enzyme digestion
