Author/Authors :
tati, khadijeh shiraz university of medical science - shiraz institute for cancer research, school of medicine - department of immunology, ايران , yazdanpanah-samani, mahsa shiraz university of medical sciences - faculty of medicine, shiraz institute for cancer research, ايران , ramezani, amin shiraz university of medical science - shiraz institute for cancer research, school of medicine, school of advanced medical sciences and technologies - department of medical biotechnology, ايران , maymand, elham mahmoudi shiraz university of medical science - shiraz institute for cancer research, school of medicine, ايران , ghaderi, abbas shiraz university of medical science - shiraz institute for cancer research, school of medicine, ايران
Abstract :
Background: CD52is a small glycoprotein with a GPI anchor at its C-terminus. CD52 is expressed by Normal and malignant T and B lymphocytes and monocytes. There are detectable amounts of soluble CD52 in plasma of patients with CLL and could be used as a tumor marker. Although the biological function of CD52 is unknown but it seems that CD52 may be involved in migration and activation of T-cells .The aim of this study was to clone and express human CD52 gene in CHO cell line and studying its function in more details. Methods:Based on GenBank databases two specific primers were designed for amplification of cd52 gene. Total RNA was extracted from Raji cell line and cDNA synthesized. Amplified fragment was cloned in pBudCE4.1 vector. The new construct was transfected to CHO-K1 cell line using electroporation method. Expression of recombinant CD52 protein was evaluated by Real time PCR and flow cytometry methods. Results:Amplification of CD52 gene using specific primers on RajicDNA showed a 209 bp band. New construct was confirmed by PCR and restriction pattern and sequence analysis. The new construct was designated as pBudKT1. RT-PCR analysis detected cd52 mRNAs in transfected cells and Flow cytometry Results showed that 78.4 % of cells represented CD52 in their surfaces. Conclusions:In conclusion, we established a human CD52 positive cell line, CHO-CD52, and the protein was expressed on the membrane. Cloning of the CD52 gene could be the first step for the production of therapeutic monoclonal antibodies and detection systems for soluble CD52 in biological fluids.
