Gene chip technique in detection of vancomycin-resistant Enterococcus gene / 军事医学

ABSTRACT

Objective To develop a detection method based on the technology of gene chips which can quickly distinguish genes of Enterococcus faecalis, E.faecium and vancomycin resistance.Methods Based on the specific gene ( ddl) sequences of two types of Enterococcus from GenBank, oligonucleotide probes which could detect and distinguish special genes and drug resistance genes ( vanA,vanB) of Enterococcus were designed and compounded.Then,the probes were dotted to modified slide.The target DNA fragments of vancomycin-resistant Enterococcus ( VRE) were labeled with biotin by multiple PCR amplification, and then hybridized with oligonucleotide probes on slide.The results were analyzed by portable imager.The multiple PCR system, hybridization reaction and condition of the chemiluminescence method were optimized before the specificity, sensitivity and reproducibility of the chip were evaluated.Results One universal primer, four specific primers, one universal probe and four specific probes were selected.This gene chip was demonstrated of high specificity and repeatability.The detection sensitivity was 103 CFU/ml.The gene chip detection results of 10 clinical samples were basically consistent with the drug sensitivity test ( 8/10 ) .Conclusion A gene chip technique for the detection of VRE is established successfully.It is possible to distinguish the type of VRE and detect the genetic phenotypes of drug resistance by gene chip technique.