[Expression of mouse lipopolysaccharide response protein LRP in E. coli and preparation of rabbit anti-mLRP serum].
OBJECTIVE
To express mouse lipopolysaccharide response protein (mLRP) and prepare rabbit anti-mLRP serum.


METHODS
The predicted mouse lrp cDNA sequence was obtained by splicing homologous ESTs by comparing human lrp cDNA with mouse ESTs. Then the primers were designed. mlrp cDNA from NIH3T3 cells stimulated with lipopolysaccharide (LPS) was amplified by RT-PCR and was cloned into prokaryotic expression vector pTAT to construct recombinant expression vector pTAT-mlrp. The His-TAT-mLRP fusion protein was expressed in E. coli BL21(DE3) and was used to immunize the rabbits to get rabbit anti-mLRP serum. The anti-serum was purified by the acetone precipitation method. The specificity of the rabbit anti-mLRP serum was determined by Western blot.


RESULTS
The predicted length of mlrp cDNA was 1905 bp. The encoding region of the cloned mlrp cDNA, 1554 bp, was inserted into pTAT. The His-TAT-mLRP fusion protein was expressed successfully in E. coli. The rabbit anti-mLRP serum was prepared by immunizing the rabbit with mLRP protein.


CONCLUSIONS
The successful expression of mLRP and the preparation of rabbit anti-mLRP serum lays the foundation for further study of the function of mLRP.
Dai
Zhong-ming
ZM
Department of Biochemistry and Molecular Biology, Fourth Military Medical University, Xi'an 710032, China. daizmly@yahoo.com
Chen
Su-min
SM
Du
Ke-jun
KJ
Chen
Nan-chun
NC
Song
Qing-he
QH
Luo
Wen-jing
WJ
Chen
Jing-yuan
JY
D000209:Acute-Phase Proteins / Q000235:genetics / Q000276:immunology* / Q000378:metabolism; D000818:Animals; D000917:Antibody Formation / Q000276:immunology*; D002352:Carrier Proteins / Q000235:genetics / Q000276:immunology* / Q000378:metabolism; D003001:Cloning, Molecular; D004926:Escherichia coli / Q000235:genetics*; D005260:Female; D015870:Gene Expression; D005822:Genetic Vectors; D008070:Lipopolysaccharides / Q000737:chemistry; D008562:Membrane Glycoproteins / Q000235:genetics / Q000276:immunology* / Q000378:metabolism; D051379:Mice; D041681:NIH 3T3 Cells; D011817:Rabbits; D011994:Recombinant Proteins / Q000235:genetics / Q000276:immunology* / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D000099231:Lipopolysaccharide-Binding Protein
D000209:Acute-Phase Proteins; D002352:Carrier Proteins; D008070:Lipopolysaccharides; D008562:Membrane Glycoproteins; D011994:Recombinant Proteins; D000099231:Lipopolysaccharide-Binding Protein