Clone,expression and identification of human recombinant IL-4 / 重庆医学

Objective Construction of human IL‐4 recombinant expression vector and then conduct the expression ,purification and identification of human recombinant IL‐4 .Methods the open reading frame of IL‐4 was amplified by nest PCR with total RNA from PBMC of healthy volunteer .And then the amplified IL‐4 was inserted into pET101/D‐TOPO ,transformed into BL21 ,ex‐pressed ,purified and indentified .Results The size of amplified open reading frame of IL‐4 was about 460 bp and the sequence was correct .After transformed into BL21 ,the IL‐4 clone with higher expression level was selected by selection of different clones insert‐ed with IL‐4 and the size of expressed ,purified IL‐4 was about 28 × 103 .Western blot results showed that the size of single band was identical with the expected protein .Conclusion Human IL‐4 recombinant protein was got successfully .

Construction of the recombinent retroviral vector rRV-hIL-4 and expression of rheumatoid arthritis in vitro / 中国实用内科杂志

Objective To construct recombinent retroviral vectors containsins the expression sequence of hIL-4,and detect the expression of target genes.Methods Design the primer with restrictive enzyme spot and amplify target gene by PCR from plasmid including hIL-4 gene.Clone the target gene into retroviral vector pLXSN,and identify the aquired plasmid by sequencing.Transfect human synoviocytes with acquired virus in vitro.Detect the protein expresssion by western blotting.Results We successfully constructed the recombinant retrovirus-rRV-hIL-4.The hIL-4 gene was transduced into human synoviocytes by recombinant retrovirus in vitro.The protein expression of genes was detected by Western blotting.Conclusion Recombinant retrovirus rRV-hIL-4 is constructed successfully.The hIL-4 gene is transduced successfully into the human synoviocytes in vitro and the transduced synoviocytes can express hIL-4 protein.