Rapid Detection of Staphylococcus aureus Based on Visual Loop-Mediated Isothermal Amplification Method

Abstract

Abstract: In this paper, a simple, rapid and sensitive method for the detection of Staphylococcus aureus based on color determination was established by using Loop-mediated isothermal amplification (LAMP). LAMP primers were designed by using the specific target gene SAR0395 of S. aureus, and the visualized LAMP reaction system was built. The optimized condition was as follows: 1.6 μmol/L inner primers ( FIP and BIP), 0.2 μmol/L outer primers (F3 and B3), 4.0 mmol/L MgSO4, 1.0 mmol/L dNTPs, 0.4 U/μL Bst 2.0 WarmStart? DNA polymerase, 120 μmol/L hydroxynaphthol blue and amplified at 60 ℃ for 60 min. Two loop primers (LF and LB) added to the visualized LAMP system can reduce the reaction time to 20 min. After 60 minutes of LAMP reaction, the genomic sensitivity of the visualized LAMP assay can reach 0.010 3 fg/μL, and the lowest content of colonies can reach 1.9 CFU/mL. The specificity and reliability of the visualized LAMP assay was confirmed by using 46 S. aureus and 24 non-S. aureus strains. The visualized LAMP assay established in this study provides a new technique for the rapid detection of S. aureus.

Key words: visualization, loop-mediated isothermal amplification (LAMP), detection, Staphylococcus aureus, hydroxynaphthol blue

CLC Number:

TS207.4

References

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