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Intranasal Administration of Recombinant Influenza Vaccines in Chimeric Mouse Models to Study Mucosal Immunity
Published on: June 25, 2015
[Construction and expression of regulatable recombinant adenovirus vector for mouse PLP-Ig chimeric protein]
Yan Zhang1, Meng-lei Liu, Qian Shen
1Department of Laboratory diagnosis, Changhai Hospital, Second Military Medical University, Shanghai 200433, China. zhangyansh@sina100.com
Aim:
To construct a regulatable recombinant adenovirus vector expressing mouse proteolipid protein (PLP)-Ig.
Methods:
Total RNA was extracted from WT-1 hybridoma cells to clone Ig heavy chain Fc fragment gene. PCR was carried out to link PLP(139-151) gene and signal peptide with Fc fragment gene. pTRE-shuttle vector was used to ligate PLP-Ig gene and the backbone of the replication-incompetent adenoviral vector. After confirming the desired recombinant adenovirus by PCR and restriction endonuclease digestion analysis, it was packaged and propagated in HEK 293 cells. Tetracycline was used to regulate the expression of PLP-Ig, and Western blot was used to detect the gene expression.
Results:
The expression of PLP-Ig was confirmed by Western blot and its expression could be regulated by tetracycline.
Conclusion:
The regulatable recombinant adenovirus vector for mouse PLP-Ig was constructed and expressed successfully, which lays the foundation for further study on gene therapy of EAE and induction of immune tolerance.
