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A new method for multilocus enzyme electrophoresis, based on electrophoretic transfers to nitrocellulose after polyacrylamide-agarose gel electrophoresis was explored. Electrophoretic separation was performed on 1-mm-thick slab gels with 6-mul samples of bacterial extracts and was followed by serial 5-min consecutive transfers. The transferability of 19 metabolic enzymes of Klebsiella strains was studied and allowed the simultaneous examination of one enzyme in the separation gel and at least five enzymes on nitrocellulose sheets. The resolution of enzyme bands was increased on nitrocellulose; thus, well-separated bands were recorded for nucleoside phosphorylase, peptidase, and phosphoglucose isomerase whereas their mobility variants could not be clearly distinguished in the separation gel because of stain diffusion. The study of genetic relationships of 42 strains of Klebsiella pneumoniae and 24 strains of Klebsiella oxytoca demonstrated the reliability of the method, since clustering analysis of electrophoretic types, based on electrophoretic polymorphism of 10 metabolic enzymes, showed two main clusters well correlated with the two species. The 57 electrophoretic types described confirm the usefulness of the method for the study of genetic relationships between closely related strains.

applied and environmental microbiology

Electrophoretic Transfer from Polyacrylamide Gel to Nitrocellulose Sheets, a New Method To Characterize Multilocus Enzyme Genotypes of Klebsiella Strains