For COVID-19 vaccine updates, please review our information guide. For patient eligibility and scheduling availability, please visit VaccineTogetherNY.org.

Highly efficient modification of bacterial artificial chromosomes (BACs) using novel shuttle vectors containing the R6Kgamma origin of replication.

TitleHighly efficient modification of bacterial artificial chromosomes (BACs) using novel shuttle vectors containing the R6Kgamma origin of replication.
Publication TypeJournal Article
Year of Publication2002
AuthorsGong S, Yang XWilliam, Li C, Heintz N
JournalGenome Res
Volume12
Issue12
Pagination1992-8
Date Published2002 Dec
ISSN1088-9051
KeywordsAnimals, Chromosomes, Artificial, Bacterial, DNA Probes, DNA, Bacterial, Escherichia coli, Gene Library, Genetic Vectors, Hexosyltransferases, Humans, Mice, Mice, Transgenic, Mutagenesis, Insertional, Rec A Recombinases, Replication Origin
Abstract

Bacterial artificial chromosome (BAC) mediated transgenesis has proven to be a highly reliable way to obtain accurate transgene expression for in vivo studies of gene expression and function. A rate-limiting step in use of this technology to characterize large numbers of genes has been the process with which BACs can be modified by homologous recombination in Escherichia coli. We report here a highly efficient method for modifying BACs by using a novel set of shuttle vectors that contain the R6Kgamma origin for DNA replication, the E. coli RecA gene for recombination, and the SacB gene for negative selection. These new vectors greatly increased the ease with which one can clone the shuttle vectors, as well as screen for co-integrated and resolved clones. Furthermore, we simplify the shuttle vector cloning to one step by incorporation of a "built-in" resolution cassette for rapid removal of the unwanted vector sequences. This new system has been used to modify a dozen BACs. It is well suited for efficient production of modified BACs for use in a variety of in vivo studies.

DOI10.1101/gr.476202
Alternate JournalGenome Res.
PubMed ID12466304
PubMed Central IDPMC187570
Grant ListN01-NS0233 / NS / NINDS NIH HHS / United States
NS 39636-03 / NS / NINDS NIH HHS / United States