Submitted to: Journal of Virological Methods
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 10/30/2006
Publication Date: 12/4/2006
Citation: Slack, J.M., Lihoradova, O., Ogay, I., Abdukarimov, A., Lynn, D.E. 2006. The homingbac baculovirus cloning system: an alternative way to introduce foreign dna into baculovirus genomes without bacmids or homologous recombination. Journal of Virological Methods. 140(1-2):59-65.
Interpretive Summary: Insect viruses have been used for over 25 years for producing foreign proteins from a wide variety of sources. These proteins have been used in a variety of research, human and veterinary medicine, and pest control applications. The most commonly used techniques for inserting the gene for the foreign protein into the insect virus can require considerable effort to obtain a stable recombinant virus capable of producing a correct, functional gene product. The present study uses a different approach for inserting the gene into the virus that simplifies the process and yields a more stable modified virus. We call our cloning system the Homingbac system because it is relies on homing endonucleases (enzymes that cut DNA at specific sites). This technique has been demonstrated in a variety of insect virus systems and has potential in both expression vector (for producing proteins) and biopesticide applications. Scientists and companies using insect virus or other expression vector systems can use this information.
Technical Abstract: Most recombinant baculovirus cloning systems use technologies involving either homologous recombination or bacmid transposition. These systems require foreign genes to be cloned into bacterial plasmid vectors prior to introduction into baculovirus genomes. We have developed a more efficient baculovirus cloning system that allows direct cloning of foreign genes into baculoviruses. Our cloning system combines restriction endonucleases with homing endonucleases. We call this system the “Homingbac system” and demonstrate the direct cloning of a polymerase chain reaction-amplified b-glucuronidase gene cassette into a baculovirus genome. Our Homingbac system was also engineered into the group I and group II nucleopolyhedroviruses. This is the first time a common baculovirus cloning system has been made for multiple baculovirus species from group I and group II NPVs.