AdHTS: a high-throughput system for generating recombinant adenoviruses.
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.
R&D Center, BRN Science, Golden Helix, Seoul National University, Seoul 151-742, Republic of Korea.
Eun-Wook
EW
Choi
Dong-Seung
DS
Seen
Yong Bhum
YB
Song
Ho-Sun
HS
Son
Neon-Cheol
NC
Jung
Won-Ki
WK
Huh
Ji-Sook
JS
Hahn
Kyungjin
K
Kim
Jae-Yeon
JY
Jeong
Tae Gyu
TG
Lee
D000256:Adenoviridae / Q000235:genetics*; D001709:Biotechnology / Q000379:methods; D003001:Cloning, Molecular / Q000379:methods*; D004279:DNA, Viral / Q000235:genetics; D005822:Genetic Vectors; D057809:HEK293 Cells; D057166:High-Throughput Screening Assays / Q000379:methods*; D006801:Humans; D011995:Recombination, Genetic
D004279:DNA, Viral