Volume 14, Issue 3 (11-2000)                   Med J Islam Repub Iran 2000 | Back to browse issues page

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ASADI KARAM G, KOOTEN C, RASAEE M, DAHA M, ZANDBERGEN G, ASGHAR S. EXPRESSION OF HUMAN PROTEINASE 3 IN CHINESE HAMSTER OVARY CELLS (CHO-CELLS). Med J Islam Repub Iran 2000; 14 (3) :271-276
URL: http://mjiri.iums.ac.ir/article-1-864-en.html
EXPRESSION OF HUMAN PROTEINASE 3 IN CHINESE HAMSTER OVARY CELLS (CHO-CELLS)
GR ASADI KARAM , CV KOOTEN , MJ RASAEE , MR DAHA , GV ZANDBERGEN , SS ASGHAR
From the Department of Biochemistry, School of Medical Sciences, Tarbiat Modarres University, Tehran, I.R. Iran
Abstract:   (4769 Views)
Proteinase 3(PR3) is a human polymorphonuclear leukocyte serine proteinase and is the main target antigen for antineutrophil cytoplasmic antibodies (ANCA) found in Wegener's granulomatosis (WG). We developed a stable expression system for conformationally intact recombinant PR3 (rPR3) in Chinese hamster ovary cells (CHO-cells). The part of PR3 cDNA that encoded the active form of PR3 was selected by using appropriate primers, and a signal sequence was also added in front ofPR3 eDNA. The signal sequencePR3 (S-PR3) was cloned into the pME 18 expression vector and the result product was electroporated into E. coli (DH5 a strain). After isolation and purification, the presence of pMEI8-S-PR3 was confirmed by using appropriate restriction endonuclease and agarose gel electrophoresis. The pMEI8- S-PR3 was electroporated with CHO-cells and the presence of rPR3 was tested in culture medium after 10 days. There was 12 ng/mL rPR3 in culture medium that had activity and was recognized by ANCA in ELISA.
Keywords: Proteinase 3, ANCA, Expression system, CHO-cell.
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Type of Study: Original Research | Subject: Biological Sciences
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