芯片电泳在免核酸提取PCR产物分析中的应用

Application of chip electrophoresis in extraction of PCR products with free nucleic acid extraction reagents

目的探讨两种免核酸提取试剂联合应用DNA芯片电泳是否能快速、准确地用于新德里金属酰胺酶(NDM-1)阳性细菌的鉴定,了解该联合检测方法在流行病病原体快速鉴定中的应用价值,为流行性病原体的快速鉴定提供便捷手段。方法取1个(或0.1个)麦氏单位NDM-1(+)菌液加入免核酸提取试剂后直接进行PCR扩增,扩增产物用自制的芯片电泳进行产物分析,通过样品迁移时间与标准品比较精确计算出目的基因片段长度并大致推算产物的含量。结果两种免核酸提取试剂联合芯片电泳分析技术能快速、准确、高效地扩增出新德里金属酰胺酶NDM-1(+)细菌的目的基因片段,尤其湖南某厂家的试剂灵敏度更高。结论该联合检测方法在流行病尤其是突发性病原体的快速鉴定、监测中具有潜在的临床应用价值。

OBJECTIVE To explore the two kinds of free nucleic acid extraction reagents combined with DNA chip electrophoresis in rapid and accurate identification of New Delhi metallo-lactamase（NDM-1）-positive strains and understand the value of this combination method in rapid identification of prevalent pathogens so as to offer convenient means for rapid identification of prevalent pathogens.METHODS One（or 0.1）McNamara units NDM-1（＋）bacterium was added to free nucleic acid extraction reagents for direct PCR amplification,then the amplified products were analyzed by means of the chip electrophoresis,and the lengths of gene fragments were accurately calculated through the migration time of the samples and comparison with the standard so as to roughly calculate the contents of the products.RESULTS The two kinds of free nucleic acid extraction reagents combined with chip electrophoresis could amplify the targeted gene fragments in the NDM-1（＋）-producing bacteria,and the reagent produced by a company in Human,in particularly,had higher sensitivity.CONCLUSION The combination detection method has potential clinical value in rapid identification and surveillance of epidemic,especially the sudden pathogens.