一种适用于乳制品基因组DNA快速提取方法的研究

目的对比NaOH裂解法、PBS裂解法以及直接煮沸法3种方法提取乳制品中核酸的提取效果,优化提取条件,确定一种更适用于现场检测、简便快速的的乳制品DNA快速提取技术。方法以牛奶、水牛奶、牦牛奶、羊奶、骆驼奶、以及驴奶6种常见的乳制品为材料,分别用NaOH裂解法、PBS裂解法以及直接煮沸法3种提取方法提取乳制品中的DNA,并根据裂解液用量和裂解时间进行优化,通过PCR扩增和琼脂糖凝胶电泳分析,检测DNA提取的质量和灵敏度。结果 NaOH裂解法能够提取所有物种的乳制品DNA,而且可以在最佳裂解条件下提取模拟掺假混合乳的DNA进行检测,发现其检测限能达到1%的牛奶含量。结论该方法取样量小,成本低,在15 min内即可完成快速提取,为实验室乳制品DNA定性或定量鉴别,以及乳制品的现场掺假鉴别提供了一种快速灵敏低成本的样品前处理技术。

Study on rapid extraction of genomic DNA from dairy products

Objective To compare the extraction effect of nucleic acid from dairy products by NaOH lysis method, PBS lysis method and direct boiling method, optimize the extraction conditions, and determine a fast and convenient milk DNA extraction technology, which was suitable for on-site detection. Methods With 6 common dairy products such as cow milk, water buffalo milk, yak milk, goat milk, camel milk, and donkey milk, NaOH lysis method, PBS lysis method and directly boiling method were used to extract DNA from dairy products, and optimized the dosage of cracking and cracking time, then analyzed the quality of the DNA and sensitivity of the method by PCR amplification and agarose gel electrophoresis analysis. Results The NaOH lysis method could extract DNA from dairy products of all species, and the limit of detection could reach to 1% cow milk by extracting DNA from simulated adulterated mixed milk under the optimal cracking conditions. Conclusion This study proposes a low-cost and little-sampling DNA extraction of dairy products, which can be quickly finished within 15 min, and provide a rapid, sensitive and low-cost sample pretreatment technology for the qualitative or quantitative detection of dairy products on-site.