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CONSTRUCTION AND TRANSFECTION OF HUMAN MUTANT CD59 EUKARYOTIC EXPRESSION SYSTEM

REN SHU-RONG,GAO MEI-HUA,WANG QIU-BO, et al(Department of Immunology,Qingdao University Medical College,Qingdao 266021,China)  
Objective To construct a human mutant CD59(HM3) eukaryotic expression system and transfect it into CHO so as to get a stable-transfection cell clone.MethodsHM3 DNA was performed by recombinant PCR using human CD59 cDNA template and designed mutagenic primers.HM3 DNA was inserted into the mammalian expression vector pALTER and transfected into CHO together with the selection marker pcDNA3,which conferred resistance to G418.The mutant clones were propagated in great amounts for further research.ResultsRecombinant plasmids of pALTER-HM3 were successfully constructed according to analysis of sequence and enzyme digestion.The mutant gene was about 500 bp.Stable transfectants were selected by the addition of G418.ConclusionHM3 DNA is successfully constructed.Appropriate cell dilution is useful to get purified transfected cell clones.
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