悬浮芯片系统快速检测常见葡萄球菌的初步实验研究

Experimental study on rapid detection of common Staphylococci based on liquichip system

目的建立以悬浮芯片为技术基础的临床常见葡萄球菌的快速检测方法。方法以细菌16S rRNA基因保守区序列设计1对通用引物，分别采用对称PCR和不对称PCR扩增5种葡萄球菌，经测序确定后，采用悬浮芯片系统对不同的PCR产物进行检测。结果2种PCR均扩增出5种葡萄球菌目的片段（780-820bp），不对称PCR得到了大量单链产物，其产物在悬浮芯片系统的检测信号高于对称PCR，悬浮芯片系统对5种葡萄球菌的鉴定率为100％。结论16SrRNA可以作为细菌快速鉴定的靶序列，不对称PCR产物可以显著提高与悬浮芯片杂交检测的灵敏度，悬浮芯片系统在鉴定细菌方面具有简单快速、高通量、高检出率等特点，可以作为细菌快速鉴定的一种新方法进行深入开发利用。

Objective To establish a new method for rapid detection of common Staphylococci based on liquiehip system. Methods Five species of Staphylococci were detected by symmetrical PCR and unsymmetrical PCR. A pair of general primers based on highly conserved segment of 16 S rRNA gene was designed for the above PCR. The amplified products of 5 species of Staphylococci were sequenced and detected by applying liquichip system. Results The highly conserved segments of 16 S rRNA gene from 5 species of Staphylococci were amplified by applying two different PCR methods （symmetrical PCR and unsymmetrical PCR）, and the products were 780-820 bp in length. The unsymmetrical PCR produced a great quantity of single-strand nucleotides. The hybridization signal of unsymmetrical PCR products was stronger than that of symmetrical PCR products in liquichip system. The identificating rate of liquichip system for 5 species of Staphylococci accounted for 100%. Conclusion 16 S rRNA gene may be used as the target sequence of rapid identification for bacteria. The unsymmetrical PCR products can significantly enhance the hybridization sensibility. Liquichip system is characteristic of simply, rapid, high-throughput and high-performance identification for bacteria and may be further developed as a new method of bacterial identification.