Arjun and Patrick's Research Proposal

=BoHV-4 Vaccine carrier =

Use BoHV-4 (Bovine Herpes Virus 4) as a carrier to secrete a specific non-toxic antigen from a virus known to be harmful to humans, inject it into cell, and force the immune system to make antibodies against it. Possible Viruses to Target: Human Papillomavirus Herpes HepatitisB etc. Original Paper link: Establishment of a Bovine Herpesvirus 4 based vector expressing a secreted form of the Bovine Viral Diarrhea Virus structural glycoprotein E2 for immunization purposes Summary of Article: A recombinant BoHV-4, containing a secreted form of the bovine viral diarrhea virus structural glycoproteins was created using a recombination-mediated genetic engineering approach. When rabbits and sheep were infected with this modified BoHV-4, large levels of antibodies were formed. This shows that BoHV-4 is a hopeful vector for vaccine purposes

Necessary components of expressed antigen:
Removal of the putative transmembrane domain Addition of a 14 amino acids peptide at its carboxyl terminal Immunoglobulin K signal peptide added to the amino terminal

Why are they needed?
Removal of putative transmembrane domain: Allows for the protein product to be secreted rather than inserted into membrane

Addition of 14 amino acids at carboxyl terminal: postulated and experimentally shown to increase secretion of protein; replaced transmembrane domain.

Immunoglobulin K signal peptide addition to amino terminal: Forces the immune system to recognize the secreted structure.

Why BoHV-4?
Non-pathogenic Low probability of oncogenicity Capable of carrying large amounts of foreign DNA Ability to infect several different types of cells from different species Ability to maintain transgene expression in either differentaited or undifferentiated cells

How to insert antigen into BoHV-4?
Recombination of modified antigen (as discussed before) DNA with BoHV-4 DNA, Cloned as bacterial artificial chromosome. Verify by using Kanamycin resistance gene in antigen cassette and selecting for it.

Predicted Outcomes
If all goes well, the use of BoHV-4 as a vehicle for the vaccine should allow for the cells to produce antibodies to the virus. If nothing goes according to plan, either the BoHV-4 is a bad vehicle or the antigen fails to allow the immune system to produce antibodies

Other Resources
Exploiting persistent infection for selection of bovine herpesvirus 4 recombinants

Potential of bovine herpesvirus 4 as a gene delivery vector