Cloning, prokaryotic expression, and biological analysis of recombinant chicken IFN-gamma.
The full-length chicken interferon-gamma (CHIFN-gamma) gene was amplified by reverse transcription-PCR using total RNA extracted from the spleen cells of white Leghorn chicken, a local Chinese breeding species. A truncated CHIFN-gamma gene without the N-terminal signal peptide sequence was cloned into prokaryotic expression vector pET30a, resulting in a recombinant plasmid pET-30a-CHIFN-gamma. After the recombinant plasmid was transformed into host cells BL21(DE3)pLysS, the expression of CHIFN-gamma was induced by isopropyl beta-D-thiogalactoside (IPTG). Rabbit antiserum was raised using the soluble CHIFN-gamma as immunogen. Immunoreactivities of the CHIFN-gamma and its antiserum were investigated using immunoblotting and ELISA. Moreover, the antiviral effect of the CHIFN-gamma was analyzed. Our data indicate that the CHIFN-gamma is biologically active.
Li
Guangxing
G
College of Veterinary Medicine, Northeast Agricultural University, Harbin, China.
Zeng
Yan
Y
Yin
Jiechao
J
Lillehoj
Hyun S
HS
Ren
Xiaofeng
X
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D015153:Blotting, Western; D002478:Cells, Cultured; D002645:Chickens; D002522:Chlorocebus aethiops; D003001:Cloning, Molecular; D004591:Electrophoresis, Polyacrylamide Gel; D004797:Enzyme-Linked Immunosorbent Assay; D004926:Escherichia coli / Q000378:metabolism; D005786:Gene Expression Regulation / Q000502:physiology*; D005822:Genetic Vectors; D007371:Interferon-gamma / Q000235:genetics* / Q000276:immunology; D008969:Molecular Sequence Data; D010957:Plasmids; D011817:Rabbits; D011994:Recombinant Proteins; D014709:Vero Cells
D011994:Recombinant Proteins; D007371:Interferon-gamma