改良十六烷基三甲基溴化铵-磁珠核酸提取方法及在植物转基因检测中的应用

Improved extraction method of cetyltrimethylammonium ammonium bromide-magnetic beads nucleic scid and its application in plant transgenic detection

中文关键词:  改良的十六烷基三甲基溴化铵-磁珠  基因组DNA提取  植物转基因检测

英文关键词:improved cetyltrimethylammonium ammonium bromide-magnetic beads  genomic DNA extraction  plant transgenic detection

中文摘要:

      目的 建立一种快速、高效、便捷的植物基因组DNA提取方法并运用于植物转基因检测。方法 改良传统的十六烷基三甲基溴化铵(cetyltrimethylammonium ammonium bromide, CTAB)方法, 并结合磁珠吸附基因组DNA, 配合核酸自动提取系统提取水稻和加工米粉中的核酸, 荧光聚合酶链式反应(polymerase chain reaction, PCR)检测Bt63大米转基因成分, 并与另外4种提取方法的结果进行对比, 评价提取效果。结果 5种不同方法中CTAB-磁珠法提取的样品基因组DNA浓度和质量最佳, 荧光PCR检测低浓度Bt63转基因成分(0.01%)的Ct值最小, 检测结果最好。结论 该方法可高效快速地提取植物核酸, 在低含量的转基因成分检测中相比其他几种方法具有绝对优势。

英文摘要:

      Objective To establish a rapid, efficient and convenient plant genomic DNA extraction method and apply to the detection of plant genetically modified organisms. Methods The DNA extraction method of cetyltrimethylammonium ammonium bromide (CTAB) was improved, silica hydroxyl magnetic beads was used to extract the genomic DNA, and the nucleic acids in rice and rice vermicelli were extracted by the automatic nucleic acid extraction system. The genetically modified components of Bt63 in the extracted DNAs were detected by fluorescent polymerase chain reaction (PCR), and the results were evaluated by comparing with the other 4 extraction methods. Results The concentration and quality of genomic DNA extracted by improved CTAB-magnetic beads method were the best among the 5 different methods, and the low concentration of Bt63 (0.01%) detected by fluorescence PCR had the lowest Ct value among the results which obtained from 5 different methods. Conclusion This method can extract plant nucleic acid efficiently and quickly, and has absolute advantages over other methods in the detection of transgenic components with low content.