Molecular cloning of caprine IL-6 cDNA and its expression in insect cells.
To analyze the interactions among immunoneuroendocrinological systems in a miniature Shiba goat, we attempted to clone and express caprine cDNA-encoding interleukin-6 (IL-6) which is well-known to exert a variety of effects on neuroendocrinological functions in small experimental animals. The cDNA-encoding caprine IL-6 was molecularly cloned from a lipopolysaccharide-stimulated adherent splenocyte library by the reverse transcription-polymerase chain reaction. The sequence analysis shows that the cloned caprine IL-6 cDNA encodes a precursor protein of 208 amino acids that is processed to mature protein of 180 amino acids with a predicted molecular weight of 20,524. The amino acid sequence of caprine IL-6 displays 97.8 and 51.1% degree of homology with ovine and human equivalents, respectively. A recombinant baculovirus containing the caprine IL-6 cDNA was shown to induce insect cells to efficiently secrete caprine IL-6 into culture supernatant up to 10(5) U/ml, as confirmed by Western blot analysis and bioassay with an IL-6-dependent cell line.
Takakura
H
H
Department of Veterinary Science, National Institute of Health, Shinjuku-ku, Tokyo, Japan.
Mori
Y
Y
Tatsumi
M
M
D000595:Amino Acid Sequence; D000818:Animals; D016367:Baculoviridae / Q000235:genetics; D001483:Base Sequence; D002478:Cells, Cultured; D003001:Cloning, Molecular; D015870:Gene Expression; D015723:Gene Library; D006041:Goats / Q000235:genetics*; D006801:Humans; D007313:Insecta; D015850:Interleukin-6 / Q000235:genetics* / Q000378:metabolism*; D008070:Lipopolysaccharides / Q000494:pharmacology; D008264:Macrophages; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction; D011995:Recombination, Genetic; D017422:Sequence Analysis, DNA; D017386:Sequence Homology, Amino Acid; D012756:Sheep
D015850:Interleukin-6; D008070:Lipopolysaccharides