Expression of Influenza Heamagglutinin Globular Head in Different Eukaryotic Cells

Background and Aims: Influenza (flu) is a respiratory infection in mammals and birds. It is caused by an RNA virus in the family Orthomyxoviridae. The virus is divided into three main types. Influenza virus type A is found in a wide variety of bird and mammal species and can undergo major shifts in immunological properties. Hemagglutinin (HA) is an important influenza virus surface antigen that is highly topical in influenza research. In the present study, the gene encoding HA1 protein which includes Hemagglutinin globular head from influenza virus A/Tehran/18/2010 (H1N1) was cloned into a eukaryotic expression plasmid (pCDNA3) and its expression was evaluated in eukaryotic cells. Materials and Methods: HA1 gene was incised from pFastBacTHc-HA1 by digestion, purified and subcloned into eukaryotic expression vector (pCDNA3). After verification of the cloning fidelity, the recombinant plasmid was transfected into COS-7 and BHK-21 cells, and its expression was detected by RT-PCR. Results: Restriction endonuclease digestion analysis, colony PCR and DNA sequencing indicated that the recombinant plasmid pCDNA3-HA1 had been constructed successfully. After transfection into eukaryotic cells, the presence of mRNA transcripts was verified by reverse transcriptase-polymerase chain reaction (RT-PCR). Conclusion: This study is a demonstrated success in the construction of eukaryotic expression plasmid for HA1 thus providing a basis for further probing into mechanism of virus infection and exploring DNA vaccine.

[1]  J. Crowe,et al.  A Broadly Neutralizing Human Monoclonal Antibody That Recognizes a Conserved, Novel Epitope on the Globular Head of the Influenza H1N1 Virus Hemagglutinin , 2011, Journal of Virology.

[2]  Charles J. Russell,et al.  An Influenza A/H1N1/2009 Hemagglutinin Vaccine Produced in Escherichia coli , 2010, PloS one.

[3]  K. Uberla,et al.  Codon-optimization of the hemagglutinin gene from the novel swine origin H1N1 influenza virus has differential effects on CD4(+) T-cell responses and immune effector mechanisms following DNA electroporation in mice. , 2010, Vaccine.

[4]  J. Haynes,et al.  Epidermal DNA vaccine for influenza is immunogenic in humans. , 2006, Vaccine.

[5]  J. Ulmer.  Influenza DNA vaccines. , 2002, Vaccine.

[6]  Ivet Bahar,et al.  Functional motions of influenza virus hemagglutinin: a structure-based analytical approach. , 2002, Biophysical journal.

[7]  D. Klinman,et al.  The regulation of DNA vaccines. , 2001, Current opinion in biotechnology.

[8]  N. Restifo,et al.  DNA and RNA-based vaccines: principles, progress and prospects. , 1999, Vaccine.

[9]  B. Wren,et al.  Nucleic acid immunization: concepts and techniques associated with third generation vaccines. , 1999, Journal of immunological methods.

[10]  L. Babiuk,et al.  Polynucleotide vaccines: potential for inducing immunity in animals. , 1999, Journal of biotechnology.

[11]  H. Davis,et al.  Plasmid DNA expression systems for the purpose of immunization. , 1997, Current opinion in biotechnology.

[12]  H. Deng,et al.  Cytokines and costimulatory molecules as genetic adjuvants , 1997, Immunology and cell biology.

[13]  J. Ulmer,et al.  Further protection against antigenic drift of influenza virus in a ferret model by DNA vaccination. , 1997, Vaccine.

[14]  B. Dixon.  The Third Vaccine Revolution , 1995, Bio/Technology.

[15]  G. Acsadi,et al.  Direct gene transfer into mouse muscle in vivo. , 1990, Science.

[16]  M. Kheiri,et al.  Expression of antigenic determinants of the haemagglutinin large subunit of novel influenza virus in insect cells. , 2012, Folia biologica.

[17]  S. Jeon,et al.  Immunization with influenza virus hemagglutinin globular region containing the receptor-binding pocket. , 2002, Viral immunology.

[18]  R. Lamb,et al.  Orthomyxoviridae: The Viruses and Their Replication. , 1996 .