[Alignment of DNA sequences of ompL1 genes of insert fragment of recombinant plasmid, pDC38 of L. interrogans serovar lai and L. kirschneri].
In a previous study a genomic library of L. interrogans serovar lai was constructed by the present authors. Hybridization analysis (In situ, dot blot, Southern blot) with the DNA fragment containing OmpL1 (alpha-32P labeled) was performed. One of positive clones designated pDC38, was analyzed with 9 restriction enzymes (EcoRI, Bam HI, Hind III, Bgl, XbalI, ScaI, KpnI, PstI, Dra II). DNA hybridization was applied to analyze the homology of the recombinant fragment of ompL1 with the DNA of 18 strains of L. interrogans. The results showed that the homology of fragments of ompL1 were present in pathogenic Leptospira strains, but they were not in the non-pathogenic Leptospira biflexa strain Patoc I, Leptonema illini strain 3055. Therefore, Dr. David A. Haake performed the sequencing of pDC38. The results showed that pDC38 contained two inserts of 2.7 kb and 3.0 kb. The 3.0 insert contained a complete copy of the ompL1 gene. Dr. David A. Haake amplified the ompL1 gene using PCR primers specific for the ends of the gene and cloned the amplicon into pBluescript KS for sequencing. The alignment of complete DNA sequences of ompL1 of L. kirschneri and L. interrogans serovar lai showed the similarity of nucleotide sequences was 90%, variation was 10%. The derived amino acid sequence showed there was a high degree of amino acid sequence homology with ompL1 of L. kirschneri. These findings indicated that ompL1 gene was one of the important outer membrane protein genes in the L. interrogans serovar lai and using the probe pDC38 might provide a good tool for classification and identification of Leptospira.
Dai
B
B
Research Unit of Leptospirosis, School of Basic Medical Sciences, WCUMS, Chengdu 610041.
Chen
Z
Z
Haake
D A
DA
You
Z
Z
Fang
Z
Z
D000595:Amino Acid Sequence; D001425:Bacterial Outer Membrane Proteins / Q000235:genetics*; D001483:Base Sequence; D004269:DNA, Bacterial / Q000235:genetics; D007919:Leptospira / Q000145:classification / Q000235:genetics* / Q000302:isolation & purification; D007921:Leptospira interrogans / Q000235:genetics* / Q000302:isolation & purification; D008969:Molecular Sequence Data; D010957:Plasmids / Q000235:genetics; D011994:Recombinant Proteins / Q000235:genetics; D016415:Sequence Alignment; D017422:Sequence Analysis, DNA
D001425:Bacterial Outer Membrane Proteins; D004269:DNA, Bacterial; D011994:Recombinant Proteins; C081904:ompL1 protein, Leptospira