Construction of pLNCX/anti-CD20scFv/IgGFc/CD80/CD28/ζ eukaryotic expression vector and expression in NIH 3T3 cells |
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Authors: | HU Yong-xian YU Kang TAN Ying-xia WU Jian-bo SHEN Zhi-jian QIAN Hong-lan LIANG Bin SHAN Da-ming |
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Institution: | 1.Department of Hematology,2 Institute of Medicine,The First Affiliated Hospital,Wenzhou Medical College,Wenzhou 325000,China;3.Clinical Laboratory,Bio-Rad Laboratories Incorporation,California,USA.E-mail:yukang@wz.zj.cn |
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Abstract: | AIM: To construct a recombinant eukaryotic expression vector pLNCX/anti-CD20scFv/IgGFc/CD80/CD28/ζ and detect its expression in NIH 3T3 cells.METHODS: CD28-ζ cDNA was amplified from the plasmids pBULLET and inserted into pLNCX vector that contained anti-CD20 scFv/IgGFc/CD80 gene.The recombinant plasmids were transfected into NIH 3T3 cells,and resistant clones were obtained by G418 selection.The gene expression of the fusion protein was determined by RT-PCR and FACS.RESULTS: The recombinant eukaryotic vector was constructed successfully,determined by PCR and enzyme digestion analysis.The target gene was amplified from NIH 3T3 cells transfected with the vectors by RT-PCR.The FACS showed that recombinant protein was expressed in NIH 3T3 cells.CONCLUSION: Construction of pLNCX/anti-CD20scFv/IgGFc/CD80/CD28/ζ expression vector and its expression in NIH 3T3 cells lay the foundation for further research of generation of modified T lymphocytes to CD20 positive lymphoma. |
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Keywords: | Antibodies Neoplasms Gene therapy Gene expression NIH 3T3 cells |
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