[Prokaryotic expression of trigeminy artificial fusion gene of Leptospira interrogans and the immunogenicity of its products].
To construct lipL32/1-lipL21-OmpL1/2 fusion gene of Leptospira interrogans and its prokaryotic expression system, and to identify the immunogenicity of its products.



PCR using linking primers was applied to construct lipL32/1-lipL21-OmpL1/2 fusion gene and a prokaryotic expression system of the fusion gene was then established using routine genetic engineering technique. SDS-PAGE was used to examine output of the target recombinant protein rLipL32/1-LipL21-OmpL1/2. Double immunodiffusion and Western Blot assay were applied to identify immunogenicity of rLipL32/1-LipL21-OmpL1/2.



lipL32/1-lipL21-OmpL1/2 fusion gene with correct sequence and its prokaryotic expression system E.coli BL21DE3pET42a-lipL32/1-lipL21-ompL1/2 was obtained in this study. The output of rLipL32/1-LipL21- OmpL1/2 after optimisation was 37.78 mg/L. The immunodiffusion titer of rabbit antiserum against rLipL32/1-LipL21-OmpL1/2 was 1:4. The rLipL32/1-LipL21-OmpL1/2 antiserum was able to recognize rLipL32/1-LipL21-OmpL1/2, rLipL32/1, rLipL21 and rOmpL1/2. Positive Western hybridization signals were found among rLipL32/1-LipL21-OmpL1/2 and rabbit antiserum against whole cell of strain 56601 and serum from patients infected with L.interrogans serogroups Icterohaemorrhagiae, Grippotyphosa, Autumnalis and Pomona.



The fusion gene lipL32/1-lipL21-OmpL1/2 and its prokaryotic expression system were successfully constructed in this study. The expressed fusion protein can be used as the antigen for developing universal genetic engineering vaccine and universal serological tests of leptospirosis.
Luo
Dong-jiao
DJ
Faculty of Basic Medicine, Qian-jiang College of Hangzhou Normal University, Hangzhou 310013, China.
Qiu
Xiao-feng
XF
Wang
Jiang
J
Yan
Jin
J
Wang
Hai-bin
HB
Zhou
Jin-cheng
JC
Yan
Jie
J
D000818:Animals; D000942:Antigens, Bacterial / Q000096:biosynthesis* / Q000235:genetics; D001425:Bacterial Outer Membrane Proteins / Q000096:biosynthesis* / Q000235:genetics; D001428:Bacterial Vaccines / Q000276:immunology; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D006801:Humans; D007921:Leptospira interrogans / Q000235:genetics* / Q000276:immunology; D008074:Lipoproteins / Q000096:biosynthesis* / Q000235:genetics; D011817:Rabbits; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis / Q000235:genetics / Q000276:immunology*; D014614:Vaccines, Synthetic / Q000276:immunology
D000942:Antigens, Bacterial; D001425:Bacterial Outer Membrane Proteins; D001428:Bacterial Vaccines; C545585:LipL21 protein, Leptospira interrogans; C406589:LipL32 protein, Leptospira; D008074:Lipoproteins; D011993:Recombinant Fusion Proteins; D014614:Vaccines, Synthetic; C081904:ompL1 protein, Leptospira