Cloning and expression of goat interleukin-18 gene.
We isolated and sequenced a 480 bp cDNA encoding mature goat interleukin-18 (gIL-18) from alveolar macrophages and splenocytes activated with LPS by RT-PCR. The gIL-18 gene was cloned into pET32a (+) vectors and sequenced. Nucleotide sequence of gIL-18 shares high homology with cattle. Fusional expression with pET32a (+) of gIL-18 of about 38kD was obtained by SDS-PAGE analysis after induction by IPTG in the E. Coli BL21 expression system. The recombinant protein can induce IFN-gamma production in PBMC. The IL-18 mRNA was constitutively detected in goat alveolar macrophages with or without LPS, While, enhanced expression was detected in splenocytes and liver cells if treated by LPS, and can be weakly detected in Peripheral blood mononuclear cells (PBMCs) treated by activators. Significant deference of IL-18 mRNA level may reflect the capacity to produce mature IL-18 in such tissues.
Liu
Wen-qiang
WQ
College of Veterinary Medicine, Shandong Agricultural University, 61 Daizong Street, Taian, Shandong, 271018, China.
Zhao
Hong-kun
HK
Gao
Yun-dong
YD
Zhong
Ji-feng
JF
D000818:Animals; D001483:Base Sequence; D003001:Cloning, Molecular; D017931:DNA Primers; D018076:DNA, Complementary / Q000235:genetics; D004591:Electrophoresis, Polyacrylamide Gel / Q000662:veterinary; D015870:Gene Expression; D006041:Goats / Q000235:genetics*; D007371:Interferon-gamma / Q000378:metabolism; D020382:Interleukin-18 / Q000235:genetics* / Q000378:metabolism; D007963:Leukocytes, Mononuclear / Q000378:metabolism; D008070:Lipopolysaccharides / Q000378:metabolism; D008099:Liver / Q000378:metabolism; D016676:Macrophages, Alveolar / Q000378:metabolism; D008969:Molecular Sequence Data; D012333:RNA, Messenger / Q000378:metabolism*; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000662:veterinary; D017422:Sequence Analysis, DNA / Q000662:veterinary; D013154:Spleen / Q000378:metabolism
D017931:DNA Primers; D018076:DNA, Complementary; D020382:Interleukin-18; D008070:Lipopolysaccharides; D012333:RNA, Messenger; D007371:Interferon-gamma