[The construction and expression of eukaryotic expression plasmid of Legionella pneumophila immunogenic protein].
OBJECTIVE
To construct the recombinant plasmid of Legionella pneumophila immunogenic protein gene (ip) and detect the immunoprotein expression in NIH3T3 cells.


METHODS
The immunogenic protein gene was amplified from DNA of Legionella pneumophila by polymerase chain reaction (PCR), then inserted into pcDNA3. 1 (+) vector. The recombinant plasmid was named as pcDNA3. 1-ip and analyzed with restriction-endonuclease Hind III and BamH I digestion, PCR and DNA sequencing techniques. The NIH3T3 cell was transfected by recombinant plasmid pcDNA3. 1-ip with lipofection strategy. The transient and stable expression products of immunogenic protein gene were detected by immunofluorescence and Western blot.


RESULTS
We have successfully constructed the recombinant plasmid of Legionella pneumophila immunogenic protein gene. It was found that there was dark green fluorescence on the cell membrane and inside the cell. Our results showed that NIH3T3 cell was transfected by pcDNA3. 1-ip successfully. The rabbit serum antibody of Legionella pneumophila detected the NIH3T3 cell transfected stably by pcDNA3. 1-ip to express the immunogenic protein with the relative molecular mass 29 X 1093).


CONCLUSIONS
We have successfully expressed immunogenic protein of Legionella pneumophila. The expressed product showed the good immunogenicity and immunoreactivity.
Yang
Zhi-Wei
ZW
Department of Parasiotology, West China School of Preclinical and Forensic Medicine, Sichuan University, Chengdu 610041, China.
Chen
Jian-Ping
JP
Liu
Ming-Fie
MF
Zhang
Lei
L
Liu
De-Song
DS
D000818:Animals; D001426:Bacterial Proteins / Q000235:genetics* / Q000276:immunology; D015153:Blotting, Western; D015870:Gene Expression; D007162:Immunoproteins / Q000235:genetics* / Q000276:immunology; D016952:Legionella pneumophila / Q000235:genetics*; D051379:Mice; D041681:NIH 3T3 Cells; D010957:Plasmids / Q000235:genetics*; D016133:Polymerase Chain Reaction; D011817:Rabbits; D015183:Restriction Mapping; D017422:Sequence Analysis, DNA
D001426:Bacterial Proteins; D007162:Immunoproteins