Establishment of Muti-channel Taqman-Probe Fluorescence Quantitative PCR Identification MRSA Method
10.3969/j.issn.1671-7414.2016.03.007
- VernacularTitle:多通道Taqman-探针荧光定量PCR鉴定MRSA方法的建立
- Author:
Changguo CHEN
;
Yanjun LI
;
Jianwei GUO
;
Qiuyuan CHEN
;
Min LIU
;
Zhijia MA
;
Xiuhong HAO
;
Qiangyuan ZHAO
- Publication Type:Journal Article
- Keywords:
taqman fluorescence probe;
fluorescence quantitative PCR;
methicillin-resistant Staphylococcusaureus;
gene;
com-bined detecting
- From:
Journal of Modern Laboratory Medicine
2016;31(3):22-25
- CountryChina
- Language:Chinese
-
Abstract:
Objective To establish the method of identifying MRSA with Taqman-fluorescence quantitative PCR basing on mecA/nuc/fem B three gene combined detecting.Methods Taking the coagulase positive MRSA,which isolated from the clinical samples and confirmed by VITEK 2 compact microbial analyzer,as the research obj ect,designed mecA/nuc/fem B specific PCR primers and Taqman fluorescent probe by bio-software PrimerPremier 5 and Designer Beacon 7,FAM,HEX and ROX markers were used to label the fluorescent probe at 5’,and the end of 3’was labeled with BHQ1,detected by fluo-rescence quantitative PCR instrment.Results ①1 g/dl gel electrophoresis results showed that the primer’s specificity of mec A/nuc/fem B were good,and molecular weight of the amplification band consistent with the expected molecular weight and no non-specific amplification band.②Three genes were obtained specific amplification in a single tube single channel and single tube multiple channel detection in PCR,and the three gene amplification effect in a single tube single tube single chan-nel and multichannel PCR similar.Conclusion Successfully established a method of multi channel Taqman-probe fluores-cence quantitative PCR identification of MRSA,mec A/nuc/fem B combined detection can effectively differentiate coagulase negative and positive MRSA,improve the accuracy of identification.
