A loop-mediated isothermal amplification method for rapid detection of NDM-1 gene

摘要

A loop-mediated isothermal amplification (LAMP) method was developed for the rapid and sensitive detection of the emerging resistance gene New Delhi Metallo-β-lactamase-1 (NDM-1), with its specificity and sensitivity having been evaluated. Six primers, including a pair of outer primers, a pair of inner primers, and a pair of loop primers, were specially designed for recognizing eight distinct sequences on the target NDM-1 gene. The amplification reaction was performed within only 40 min under isothermal conditions at 65°C in a regular water bath. The LAMP assay showed good specificity and higher sensitivity than the conventional polymerase chain reaction (PCR), with a detection limit of 1 pg genomic DNA per tube of one NDM-1-positive reference strain. The detection result for the 345 clinical samples showed 100% consistence with the result by the PCR method, and three contaminated samples could be detected correctly by LAMP assays, while they could not be detected by PCR. The LAMP method reported here demonstrated a potential and valuable means for detection of the NDM-1 gene: easy, rapid, visual, specific, accurate, and sensitive, especially useful for on-the-spot investigation.