title: [Construction of recombinant retroviruses expressing anti-CD20 scFv/CD80 /CD28/zeta gene and expression in Jurkat cell line].
abstract: OBJECTIVE
  To construct recombinant retroviruses expressing anti-CD20 scFv/CD80/CD28/zeta gene and detect its expression in Jurkat cells.
  
  
  METHODS
  CD28-zetacDNA were amplified from plasmids pBULLET and inserted into pLNCX vector that contained anti-CD20 scFv/CD80 gene. The recombinant plasmids were transfected into PA 317 cells. Retroviruses were harvested from culture medium of PA 317 cells. Then NIH 3T3 were transfected with retroviruses. Objective gene expression was determined by PCR and FACS. Jurkat cells were transfected with high titer of retroviruses and resistant clones were obtained by G418 selection. Objective mRNA was determined by RT- PCR.
  
  
  RESULTS
  The recombinant eukaryotic vector was constructed successfully by PCR and enzyme digestion analysis and objective gene was amplified from NIH 3T3 cells transfected with retroviruses by PCR; FACS showed that objective protein could be expressed in NIH 3T3 cells. Objective gene was amplified from Jurkat cells transfected with retroviruses by RT-PCR.
  
  
  CONCLUSIONS
  Recombinant retrovirus expressing anti-CD20 scFv/CD80/CD28/zeta gene was successfully constructed and objective protein could be expressed in Jurkat cells.
authors.affiliation: Department of Hematology, Wenzhou Medical College Affiliated the First Hospital, Wenzhou 32500, China.
authors.forename: Yong-Xian
authors.initials: YX
authors.lastname: Hu
authors.forename: Kang
authors.initials: K
authors.lastname: Yu
authors.forename: Ying-Xia
authors.initials: YX
authors.lastname: Tan
authors.forename: Zhi-Jian
authors.initials: ZJ
authors.lastname: Shen
authors.forename: Hong-Lan
authors.initials: HL
authors.lastname: Qian
authors.forename: Bin
authors.initials: B
authors.lastname: Liang
authors.forename: Da-Ming
authors.initials: DM
authors.lastname: Shan
mesh_terms: D018951:Antigens, CD20 / Q000235:genetics / Q000378:metabolism*; D018122:B7-1 Antigen / Q000235:genetics / Q000378:metabolism*; D018106:CD28 Antigens / Q000235:genetics / Q000378:metabolism*; D005822:Genetic Vectors; D006801:Humans; D007140:Immunoglobulin Fab Fragments / Q000235:genetics / Q000378:metabolism; D007135:Immunoglobulin Variable Region / Q000235:genetics / Q000378:metabolism*; D019169:Jurkat Cells; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D012190:Retroviridae / Q000235:genetics; D013601:T-Lymphocytes / Q000378:metabolism*; D014162:Transfection
chemical_list: D018951:Antigens, CD20; D018122:B7-1 Antigen; D018106:CD28 Antigens; D007140:Immunoglobulin Fab Fragments; D007135:Immunoglobulin Variable Region; D011993:Recombinant Fusion Proteins