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AdHTS: A high-throughput system for generating recombinant adenoviruses

Paper ID Volume ID Publish Year Pages File Format Full-Text
23626 43459 2012 7 PDF Available
Title
AdHTS: A high-throughput system for generating recombinant adenoviruses
Abstract

The need for efficient high-throughput gene delivery system for mammalian cells is rapidly increasing with the growing request for functional genomics studies and drug discoveries in various physiologically relevant systems. However, plasmid-based gene delivery has limitations in transfection efficiency and available cell types. Viral vectors have great advantages over plasmid-based vectors, but construction of recombinant viruses remains to be a big hurdle for high-throughput applications. Here we demonstrate a rapid and simple high-throughput system for constructing recombinant adenoviruses which have been used as efficient gene delivery tools in mammalian systems in vitro and in vivo. By combining Gateway-based site-specific recombination with Terminal protein-coupled adenovirus vector, the adenovirus high-throughput system (AdHTS) generates multiple recombinant adenoviruses in 96-well plates simultaneously without the need for additional cloning or recombination in bacteria or mammalian cells. The AdHTS allows rapid and robust cloning and expression of genes in mammalian cells by removing shuttle vector construction, bacterial transformation, or selection and by minimizing effort in plaque isolation. By shortening the time required to convert whole cDNA library into desired viral vector constructs, the AdHTS would greatly facilitate functional genomics and proteomics studies in various mammalian systems.

► We designed a high-throughput system for generating recombinant adenoviruses. ► Gateway recombination was combined with Terminal protein-linked adenoviral vectors. ► Recombination, transfection, and virus production is conducted in 96-well plates. ► Our system allows rapid construction of recombinant adenoviruses by removing bacterial manipulation steps.

Keywords
Ad, adenovirus; AdHTS, adenovirus high-throughput system; GOI, gene of interest; ITR, inverted terminal repeat; TP, terminal protein; TPC, terminal protein complex; pfu, plaque-forming unitAdenovirus; Terminal protein; Gateway cloning; Vector; High-throug
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AdHTS: A high-throughput system for generating recombinant adenoviruses
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Publisher
Database: Elsevier - ScienceDirect
Journal: Journal of Biotechnology - Volume 162, Issues 2–3, 31 December 2012, Pages 246–252
Authors
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Subjects
Physical Sciences and Engineering Chemical Engineering Bioengineering
Get Full-Text Now
Don't Miss Today's Special Offer
Price was $35.95
You save - $31
Price after discount Only $4.95
100% Money Back Guarantee
Full-text PDF Download
Online Support
Any Questions? feel free to contact us