Real-time Fluorescence Quantitative PCR for Detecting Extended Spectrum-β-Lactamases Genotype / 天津医药
Tianjin Medical Journal
;
(12): 839-842,后插5, 2009.
Article
in Chinese
| WPRIM
| ID: wpr-601827
ABSTRACT
Objective:
To establish a rapid method to detect drug-resistance genotypes of extended spectrum-β-Lactamases (ESBLs) produced by gram negative bacillus using the real -time fluorescence quantitative PCR.Methods:
According to clinical common genotypes of ESBLs, SHV, TEM.CTX-M.OXA and their homology, 9 pairs of specific primers were designed including SHV, TEM, CTX-M-1, CTX-M-2, CTX-M-8, CTX-M-9, OXA-1, OXA-2 and OXA-10. To extract DNA template by boiling assay, and then establish and grade up SYBR GREEN I real-time fluorescence quantitative PCR reaction system, finally definite real-time fluorescence quantitative PCR method. Its precision and range of linearity were tested. With established assay 51 multi- drug resistant ESBLs- E. coli K. pneumoniae were detected and compared with improved three dimensional extract tests.Results:
Except OXA-2, 8 genotypes SHV, TEM, CTX-M-1, CTX-M-2, CTX-M-8, CTX-M-9, OXA-1 and OXA-10 were amplified by quantitative PCR from 39 ESBLs+ and 51 multi-drug resistant ESBLs-E. coli K. pneumoniae and confirmed by sequence testing. The range of linearity was 3×10~3-3×10~8 copies/mL, r =-0.994 7. Repetitive experiments showed that the average coefficient of variation between -runs was 9.6%. Comparing with three dimensional extract test, there was no significant difference (χ2 = 1.125,P> 0.05).Conclusion:
Testing drug-resistance genotypes of ESBLs with SYBR GREEN I real-time fluorescence quantitative PCR is a rapid,specific and sensitive method, which is capable of inspecting genotypes of ESBLs from clinical strains.
Full text:
Available
Index:
WPRIM (Western Pacific)
Language:
Chinese
Journal:
Tianjin Medical Journal
Year:
2009
Type:
Article
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