Viral Production Core
The Viral Production Core provides a range of molecular biological services related to gene expression, study, and manipulation. The primary focus is on the production, concentration, purification and titer of recombinant, replication-deficient lentivirus, adenovirus, and adeno-associated viruses (AAV) expressing genes of interest to the user of the Core. Recombinant viruses are an excellent means of introducing recombinant DNA into living cells, especially in whole animal applications. The inability to replicate is a safety feature that limits the delivery of the viral contents to the immediate target, preventing its spread and escape into non-target populations.
In a typical production, the user will provide the Core with a "donor" plasmid containing the construct to be incorporated into a chosen viral vector. The Core will then package that construct into the appropriate capsid, concentrate and purify the recombinant virus, and provide a measure of the yield of virus. Related services include:
- Consultation on the choice of optimal viral delivery system and approaches to the construction of an appropriate donor plasmid
- Assistance with completion of the Biosafety Application
- Provision of a variety of plasmid vectors that can be used to generate the donor plasmid
- Construction of the donor plasmid itself
- Production of the quantities of donor plasmid needed for viral production
- Certification of lack of replication competence (needed for whole animal studies).
- Subcellularly-localized fluorescent markers. We stock a variety of plasmid DNAs that produce fluorescent proteins that are targeted to specific subcellular locations or organelles for use by users needing in vivo localization markers. Site targeted (and the fluorescent tag attached) include:
- endosomes (GFP)
- ER (YFP, CFP)
- focal adhesions (GFP)
- Golgi (YFP, CFP, Blue)
- intranuclear sites (GFP, YFP, CFP)
- mitochondria (GFP)
- inner nuclear membrane (GFP)
- inner leaflet of the plasma membrane (GFP)
- We are continuing to look for additional targeted markers and would appreciate donations from anyone who can add to this list.
- Work can not be started on any project until it is approved by the Institution Biosafety Committee. The Core will need a copy of the summary statement in the application and the approval letter before beginning work.
- AAV can currently be pseudotyped with the AAV1, AAV2, AAV6, AAV8, or AAV9 coat proteins. These different coat proteins infect different tissues or cell lines to different extents.