A sulfone group-labeled TEM-DNA probe: comparison with a 32P-labeled probe in dot-hybridization

Abstract

A non-radioactive DNA probe for the TEM-type β-lactamase gene was obtained by using the ‘Chemoprobe’ system. It was used along with a 32P-labeled TEM probe to screen for TEM β-lactamase gene in 107 bacterial isolates representing 7 Gram-negative genera and previously classified as TEM-positive or negative. The DNA to be tested was extracted from these bacterial isolates by the Birnboim-Doly method and, after blotting charged bylon membranes, it was submitted to hybridization with either the TEM ‘Chemoprobe’ or the 32P-TEM probe. The TEM ‘Chrmoprobe’ could be detect as few as 25 pg specific DNA if it was used at a concentration of 5 ng per cm 2 of membrane. The results obtained by both probes were concordant in 93.5% of the entire samples. The TEM ‘Chemoprobe’ was specific since only one false positive was observed. Furthermore, it appeared at least as sensitive as the 32P-labeled TEM probe. As the dot-hybridization with the sulfone-labeled probe was sensitive, simple ans easy to perform, it will be useful for large-scale screening in clinical laboratory.