Comparison of Four Methods for Extracting the Genomic DNA of Staphylococcus aureus

Abstract

目的：比较4种方法，包括煮沸法、改良碱裂解法、磁珠法和吸附柱法提取金黄色葡萄球菌基因DNA效果，为临床金黄色葡萄球菌感染的快速检测提供技术支撑。方法：选取20株临床分离耐甲氧西林的金黄色葡萄球菌，分别使用煮沸法、改良碱裂解法、磁珠法和吸附柱法提取基因DNA。通过PCR法扩增所提取DNA模板中金黄色葡萄球菌spa基因、mecA基因和femB基因，以评估4种方法提取效果。结果：磁珠法制备的模板中，PCR法可以扩增出所有标本的spa基因、mecA基因和femB基因；吸附柱法制备的模板中，所有标本均可扩增出spa基因和mecA基因，而仅19个标本扩增出femB基因；以改良手工碱裂解法制备的模板，所有标本均可扩增出mecA基因，19标本可以扩增出spa基因和femB基因；煮沸法制备的模板中，PCR法可以分别扩增出7个标本的spa基因，8个标本的mecA基因，所有标本均未扩增出femB基因。结论：改良碱裂解法、吸附柱法和磁珠法提取金黄色葡萄球菌基因组DNA均可以用于PCR扩增，直接煮沸法提取效果较差，不适合用于临床金黄色葡萄球菌基因组DNA提取；磁珠法用时最短，更适合于大量快速自动化提取。 Objective: To compare the effects of four methods, including boiling method, improved alkali cracking method, magnetic bead method and adsorption column method, for extracting the genomic DNA of S. aureus, selecting methods with better extraction quality and more suitable for automation, to provide technical support for rapid detection of clinical S. aureus infection. Methods: 20 clinical strains of methicillin-resistant S. aureus (MRSA) were isolated and their DNA was extracted by boiling method, improved alkali cracking method, magnetic bead method and adsorption column method. The extraction effects of spa gene, mecA gene and femB gene in S. aureus were detected by polymerase chain reaction (PCR). Results: In the DNA gene template extracted by magnetic bead method, all samples detected spa genes, mecA genes and femB genes; in the DNA gene template extracted by adsorption column method, all samples detected spa genes and mecA genes, while 19 samples detected femB genes; in the DNA gene template extracted by improved alkali cracking method, all samples detected spa genes, and 19 samples detected mecA genes and femB genes; in the DNA gene template extracted by boiling method, only 7 samples and 8 samples detected spa genes, mecA genes and none of the samples detected femB genes. Conclusion: The DNA gene template extracted by improved alkali cracking method, magnetic bead method and adsorption column method can be used for PCR assay, but the direct boiling method is not suitable for clinical S. aureus genomic DNA extraction. Magnetic bead method uses the shortest time, which is more suitable for a large number of rapid automatic extraction.