Go

Vector Core Services

A A A

Our services include:

  • The design and engineering of custom lentiviral, retroviral, and adenoviral vectors for gene delivery.
  • The packaging of lentiviral, retroviral, and adenoviral particles.
  • Providing ready-made lentiviral particles that contain eGFP, luciferase, and RFPs (dsRED, mCherry, or tdTomato.
  • The packaging of lentiviral particles that encode shRNA for gene-knockdown.
  • The packaging of lentiviral particles that encode genes responsible for induced pluripotent stem cells (iPS)


New Technologies

The Vector Core welcomes inquiries about any new or advanced technologies that are of interest to your research. If necessary, the Vector Core will work with the Office of Technology Transfer to access third party materials through material transfer agreements.

The new technologies that we can now incorporate into our service include:

  • Virus-based nanoparticles (VNPs), which are used as platform technologies for diagnostic imaging.
  • Pantropic ViraSafe Universal Lentiviral Expression System developed at our hospital and licensed to Cell Biolabs, Inc (Cat# VPK-211-PAN). This system provides increased safety measures to prevent the generation of replication competent lentiviral particles, and includes several features that boost viral production and viral titer.
  • The new retroviral-based forward somatic cell genetic platform technology, SILENCE, which Dr. Banks developed at UCLA (Banks and Bradley, Nat. Methods, 4, 51; 2007). This system uses retrovirus particles to deliver a cis acting gene silencing element in a quasi-random fashion throughout the genome of tissue culture cells to identify unknown genes responsible for a phenotype of interest. Presently, this system works in hypodiploid cells such as CHO-K1 cells and in the mammalian cell line KBM7 chronic myeloid leukemia (CML) cell line with a haploid karyotype, except for chromosome 8 M (Kotecki et al., Exp. Cell Res. 252, 273; 1999).

Detailed Description of Services

 

Vector design and construction

  • Basic cloning: A single gene is to be cloned into an existing vector backbone. No more than 2 sub-cloning steps are required. No additional sequencing is required because the investigator has a complete and accurate sequence data file of the gene in its current plasmid.
  • Basic cloning w/sequencing: A single gene is to be cloned into an existing vector backbone. No more than 2 sub-cloning steps are required. However, a complete sequence of the gene in its current plasmid is not available and sequencing must be done to confirm 5' and 3' ends.
  • Basic cloning w/PCR +2 primer sequencing: A single gene is to be cloned into an existing vector backbone. The gene must be amplified by PCR and confirmed by sequencing. The gene is smaller than 600bp*.
  • Intermediate cloning: Two fragments (e.g. cDNA + promoter) are to be cloned into an existing vector backbone. No more than 4 sub-cloning steps are required. A complete sequence of the gene in its current plasmid is known and no additional sequencing is required.
  • Intermediate cloning w/sequencing: Two genes/fragments are to be cloned into an existing vector backbone. No more than 4 sub-cloning steps are required. However, a complete sequence of the genes is not provided and must be sequenced to confirm 5' and 3' ends.
  • Intermediate cloning w/PCR: Two genes/fragments are to be cloned into an existing vector backbone. One or both of the genes/fragments must be amplified by PCR and confirmed by sequencing. The gene being amplified is smaller than 600bp*. Custom cloning: A more complex cloning strategy is required. The price will be determined by the complexity and the amount of sequencing required.


Vector packaging

  • Endotoxin-free maxi-preps: Endotoxin-free plasmid yields the best transfection efficiency in virion production. Commercially available kits are used to purify plasmid. Plasmid quality and quantity are measured.
  • Viral titering will be measured by either selectable marker (G418, puromycin, etc) or by fluorescent marker (FACS).
  • In the event that vectors do not contain a selection or fluorescent marker, titration will be conducted by qPCR with virus specific primers. This service is not included in the quoted price.
  • Virions can be concentrated by either ultracentrifugation or tangential flow. Our pricing includes testing viral particles for replication competence according to guidelines and standards established by federal laboratories.