Article

[Construction of the eukaryotic recombinant vector and expression of the outer membrane protein LipL32 gene from Leptospira serovar Lai].

Department of Infection and Immunity, Sichuan University, Chengdu 610041, China.
Xi bao yu fen zi mian yi xue za zhi = Chinese journal of cellular and molecular immunology 03/2008; 24(2):109-12. pp.109-12
Source: PubMed

ABSTRACT To construct the eukaryotic experssion vector of LipL32 gene from Leptospira serovar Lai and express the recombinant plasmid in COS-7 cell.
The LipL32 gene was amplified from Leptospira strain 017 genomic DNA by PCR and cloned into pcDNA3.1, through restriction nuclease enzyme digestion. Then the recombinant plasmid was transformed into E.coli DH5alpha. After identified by nuclease digestion, PCR and sequencing analysis, the recombinant vector was transfected into COS-7 cell with lipsome. The expression of the target gene was detected by RT-PCR and Western blot.
The eukaryotic experssion vector pcDNA3.1-LipL32 was successfully constructed and stably expressed in COS-7 cell.
The eukaryotic recombinant vector of outer membrane protein LipL32 gene from Leptospira serovar Lai can be expressed in mammalian cell, which provides an experimental basis for the application of the Leptospira DNA vaccine.

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Keywords

COS-7 cell
 
E.coli DH5alpha
 
eukaryotic experssion vector
 
eukaryotic experssion vector pcDNA3.1-LipL32
 
eukaryotic recombinant vector
 
Leptospira DNA vaccine
 
Leptospira strain 017 genomic DNA
 
LipL32 gene
 
lipsome
 
nuclease digestion
 
outer membrane protein LipL32 gene
 
recombinant plasmid
 
restriction nuclease enzyme digestion
 
RT-PCR
 
sequencing analysis
 
target gene
 
Western blot
 

Bi Huang