[Cloning and expression of flagellin gene from a Chinese Borrelia burgdorferi PD91 strain].
OBJECTIVE
To study the cloning and expression of flagellin gene from Chinese Borrelia burgdorferi, PD91 strain and to evaluate the feasibility of using recombinant protein as diagnostic antigen when comparing the gene sequence with flagellin gene from North American Borrelia burgdorferi B31.


METHODS
The piece of genes coding flagellin from Chinese Borrelia burgdorferi PD91 by polymerase chain reaction (PCR) method was obtained, and constructed recombinant plasmid, before transformed into E. coli BL21 strain, and induced. The recombinant plasmid was identified with enzyme cutoff and gene sequence comparison. Efficient expression strain was selected and the expression product was analyzed with sodium amplified polymorphic-polyacrylamide gel electrophoresis (SDS-PAGE) and Western-blot method.


RESULTS
The recombinant protein (r-flagellin) expressed in host bacteria was successful. By means of western-blot assay, the immunological response showed the same antigenicity between r-flagellin and PD91 flagellin. The piece of genes coding flagellin of PD91 was 1011 bp, but when comparing with that of North American Borrelia burgdorferi it showed 94.70% homology. Homology between the sequence of amino acid of the r-flagellin and that of B31 flagellin was 95.85%.


CONCLUSIONS
Flagellin gene of Borrelia garinii of Chinese Lyme disease spirochete was successfully cloned and expressed for the first time. It was proved that the immunoreactivity of r-flagellin was the same as the natural flagellin.
Lü
Bing
B
National Institute of Commmunicable Disease Control & Prevention, Chinese Center of Disease Control and Prevention, Beijing 102206, China.
Wan
Kang-lin
KL
Hou
Xue-xia
XX
Hao
Qin
Q
Geng
Zhen
Z
D000595:Amino Acid Sequence; D001483:Base Sequence; D025065:Borrelia burgdorferi / Q000235:genetics* / Q000302:isolation & purification; D003001:Cloning, Molecular; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D005408:Flagellin / Q000096:biosynthesis* / Q000235:genetics; D006801:Humans; D008193:Lyme Disease / Q000382:microbiology*; D008969:Molecular Sequence Data; D010957:Plasmids / Q000235:genetics; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000235:genetics
D011994:Recombinant Proteins; D005408:Flagellin