Amplification and Cloning of a Gene Encoding a 41 kDa Outer Membrane Protein (LipL41) of Leptospira interrogans Serovar Canicola

Leptospirosis has been recognized as an important reemerging infectious disease caused by pathogenic Leptospira spp. A major challenge of this disease is the application of a basic research to improve diagnostic method. Outer membrane proteins of Leptospira are potential candidates that could be useful in diagnosis. Among them the lipL41 is an immunogenic protein which is present only in pathogenic serovars. In order to evaluate genetic conservation of the lipL41 gene, we cloned and sequenced this gene from Leptospira interrogans serovar Canicola.
Following the DNA extraction from the serovar, the lipL41 gene was amplified and cloned into pTZ57R/T vector and transformed into the competent E. coli (Top10). Recombinant clones were confirmed by colony PCR and DNA sequencing. The related sequences were then analyzed and compared with the sequences in the Genbank database.
PCR amplification of the lipL41 gene resulted in a 1065 bp PCR product. The PCR based on the lipL41 gene detected all the pathogenic reference serovars of the tested Leptospira spp. It was revealed that in Iran the homology of the lipL41 gene between vaccinal and clinical serovars of Canicola was 100%. It also showed >95.9% homology with other pathogenic serovars in Genbank database, which indicates genetic conservation of this gene.
Because of the conservation of lipL41 gene among different strains of Leptospira and its exclusive presence in leptospira, it was revealed that the cloned gene could be further used as a good candidate for developing diagnostic methods such as ELISA and as positive control in diagnostic PCR.