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Pipeline 4 : Engineering Vectors & Designing Clone Constructs Day 1
Successful engineering and designing of vectors and clone constructs leads to efficient production throughout the protein expression pipeline. Protein expression bottlenecks frequently arise because functional proteins are difficult to produce as they fail to express, are expressed as insoluble aggregates, or cannot be purified by standard methods. Thus, protein engineers are forced to return to the drawing board. This usually requires designing new cloning schemes including: lengthy verification and sequence analysis of the gene or protein of interest, moving a gene from one vector to another, transfecting the vector in an alternative host, re-characterizing the expressed protein, or any or all of the above. Researchers view this process as a linear pathway: make an expression clone, try it out, and if it fails, go back to the beginning and start over--an inefficient, time-consuming and expensive process. Cambridge Healthtech Institute’s Inaugural Engineering Vectors and Designing Clone Constructs continues the tradition of applying protein discovery research leading to functional products. Learn from seasoned savvy researchers as they share their real-world experiences, applications, and results. Thursday, January 14 1:15 pm Registration for Engineering Vectors & Designing Clone Constructs
ENGINEERING VECTORS 1:45 Chairperson’s Opening Remarks 1:50 Highly Efficient Production of Diverse Protein Targets from Cell Substrates Engineered with Lentiviral Vectors Boro Dropulic, Ph.D., Founder, President & Chief Scientific Officer, Lentigen Corp. Lentiviral vectors are known as the most efficient method to deliver genes stably into cells. There are several significant advantages for using this technology for protein production: (i) Quality – multiple gram levels of protein production from mammalian cells while maintaining high levels of glycosylation; (ii) Flexibility – applicable to all mammalian cell types, including adherent or suspension cells; (iii) Speed – multiple gram quantities of fully glycosylated protein within 12 weeks of sequence availability; (iv) Robust productivity – high transgene copy number per cell mitigates risk of gene silencing; and (v) Economics – high level of production in supernatant significantly decreases upstream and downstream costs; this technology is well suited for integration with disposable bioreactor systems. 2:20 Improved in vivo Gene Expression by Improved Plasmid Engineering, Formulation as well as Improved Physical Delivery in Immune Therapy and Vaccine Models David B. Weiner, Ph.D., Professor, Department of Pathology and Laboratory Medicine, University of Pennsylvania 2:50 Purification and Characterization of a Yeast Endocytic Protein Using the Yeast Expression Plasmid pEG(KT) B. Daniel Pierce, Ph.D., Department of Biology, Johns Hopkins University The purification of proteins in large amounts is required for a variety of biochemical and biophysical experiments. The yeast plasmid pEG(KT), which we have sequenced and made amenable for modification, is particularly useful for purifying proteins that require eukaryotic post-translational modifications or cause problems when bacterially expressed. Using this plasmid, we have purified the yeast endocytic scaffold protein Pan1 for biochemical analysis of its self-interactions and biophysical experiments, such as a characterization of its intrinsic tryptophan fluorescence to obtain structural information. 3:20 Sponsored Presentation (Sponsorship Opportunity Available) 3:35 Networking Refreshment Break in the Exhibit Hall 4:30 Harnessing The Proteome: The Center for Personalized Diagnostics at Arizona State Jason Steel, B.S., Research Scientist, Center for Personalized Diagnostics,The Biodesign Institute, Arizona State University We have initiated a project to create a sequence-verified collection of full-length cDNAs representing all coding regions for several organisms in a vector system that is protein expression-ready. By using a recombination-based vector system, users are able to execute the automated transfer of 1,000’s of genes into any protein expression vector overnight. We have demonstrated that it is possible to convert the full coding potential of an organism into a single repository of full-length cDNAs that can be used for all aspects 5:00 Vector-Based Strategies for Enhanced Protein Expression in the Food Grade Bacterium Lactococcus lactis David Mills, Ph.D., Professor, Viticulture & Enology, University of California Davis Lactococcus lactis is a common starter culture used in various dairy fermentations. Because of its long history of food grade use, combined with significant acid tolerance, L. lactis is currently a popular target for oral delivery of various biotherapeutic proteins. 5:30 Options for Manufacturing Technologies and a New Approach to Expressing Therapeutic Proteins in Mammalian Cells Yune Kunes, Ph.D., Associate Director, Biologics CMC, Abbott Bioresearch Center (tentative) 6:00 Close of Day ______________________________________________________________________________________________ * Separate registration required
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