[Study on expression of Streptococcus suis serotype 2 sly gene, purification and activity of its product].
OBJECTIVE
To clone and express Streptococcus suis serotype 2 (S. suis 2) sly gene for constructing an foundation on identification of S. suis 2 protective antigen.


METHODS
The sly gene was amplified from S. suis 2 clinical isolate strain 05ZYH33 genome DNA by PCR. The gene fragment was inserted into the expression vector pET-30b(+) to build pET30b-sly. When recombinant vector pET30b-sly was identified by restriction enzyme cutting and DNA sequencing as a correct one, subsequently it was transformed to E. coli Rosetta for expression under IPTG induction. The obtained fusion protein was purified by Ni-NTA affinity chromatography. The immunologic and hemolysis activity of the purified protein was proved through Western blot and hemolysis assay respectively.


RESULTS
The PCR product was around 1500 bp. The gene segment inserted into the recombinant vector was proven to be completely identical with the sly gene sequence in the total genome sequence of S. suis 2. The target protein expressed was up to 30% of the total somatic protein under IPTG induction. The protein purity reached above 80% after purification. The protein could be recognized by human serum infected with S. suis 2 and could dissolve swine erythrocytes with the Hemolytic titer as 256.


CONCLUSIONS
The expression vector pET30b-sly was successfully constructed. The target protein could be over-expressed in E. coli and possessed its biological activity after purification.
Liu
Li-Na
LN
Department of Microbiology, Third Military Medical University, Chongqing 400038, China.
Hu
Fu-Quan
FQ
Zhu
Jun-Min
JM
Zhao
Yan
Y
Pan
Qu
Q
Li
Ming
M
Tang
Jia-Qi
JQ
D000818:Animals; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism* / Q000494:pharmacology*; D015153:Blotting, Western; D002846:Chromatography, Affinity; D006461:Hemolysis / Q000187:drug effects; D006801:Humans; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism* / Q000494:pharmacology*; D017011:Streptococcus suis / Q000235:genetics / Q000378:metabolism*; D013552:Swine
D001426:Bacterial Proteins; D011994:Recombinant Proteins