[Gene cloning and expression of mouse type I soluble interleukin 1 receptor in insect cells].
Interleukin 1(IL-1), which takes part in many physiological and pathological processes, is a kind of important cytokines. Their effects are exerted via specific IL-1 receptors (IL-1R) which are present on a wide variety of different cell types. Two types of IL-1R are identified as type I receptor (IL-1RI) and type II receptor (IL-1RII). Soluble forms of IL-1R can be produced from proteolytic cleavage of the extracellular portion of the two type receptors on cell surface. In this study, mouse type I soluble IL-1R (sIL-1RI) has been cloned and expressed in insect cells Sf9 with baculovirus as vector. Total RNA from NIH/3T3 cell was extracted with guanidinium thiocyanate followed by ultra-centrifugation in cesium chloride solution. The cDNA of sIL-1RI was amplified by RT-PCR technique. The cDNA was cloned into plasmid pAcGP67B. The recombinant plasmid pAI (pAcGP67B-sIL-1RI) was cotransfected with wild type AcNPV DNA into insect cells Sf9, because of genetic exchange occurred by homologous recombination in vivo, recombinant baculovirus rAcNPV was produced. Insect cells Sf9 were infected with the purified rAcNPV and sIL-1RI gene were expressed. SDS-PAGE analyses and Blockage assay of IL-1 beta biological activity demonstrated that the recombinant sIL-1RI had biological function and could be secreted into the medium.
Zhang
Z P
ZP
Institute of Medicinal Biotechnology, Chinese Academy of Medical Scienes & Peking Union Medical College, Beijing.
Li
Y
Y
D016475:3T3 Cells; D000818:Animals; D003001:Cloning, Molecular; D051379:Mice; D017472:Receptors, Interleukin-1 / Q000096:biosynthesis* / Q000235:genetics; D011994:Recombinant Proteins / Q000096:biosynthesis*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D018411:Spodoptera
D017472:Receptors, Interleukin-1; D011994:Recombinant Proteins