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A new approach for the identification and cloning of genes: the pBACwich system using Cre/lox site-specific recombination

Choi, Sangdun and Begum, Dilara and Koshinsky, Heather and Ow, David W. and Wing, Rod A. (2000) A new approach for the identification and cloning of genes: the pBACwich system using Cre/lox site-specific recombination. Nucleic Acids Research, 28 (7). Art. No. E19. ISSN 0305-1048. https://resolver.caltech.edu/CaltechAUTHORS:20111123-113725128

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Abstract

With current plant transformation methods (Agrobacterium, biolistics and protoplast fusion), insertion of DNA into the genome occurs randomly and in many instances at multiple sites. Associated position effects, copy number differences and multigene interactions can make gene expression experiments difficult to interpret and plant phenotypes less predictable. An alternative approach to random integration of large DNA fragments into plants is to utilize one of several site-specific recombination (SSR) systems, such as Cre/lox. Cre has been shown in numerous instances to mediate lox site-specific recombination in animal and plant cells. By incor­porating the Cre/lox SSR system into a bacterial artificial chromosome (BAC) vector, a more precise evaluation of large DNA inserts for genetic complementation should be possible. Site-specific insertion of DNA into predefined sites in the genome may eliminate unwanted ‘position effects’ caused by the random integration of exogenously introduced DNA. In an effort to make the Cre/lox system an effective tool for site-directed integration of large DNAs, we constructed and tested a new vector potentially capable of integrating large DNA inserts into plant and fungal genomes. In this study, we present the construction of a new BAC vector, pBACwich, for the system and the use of this vector to demonstrate SSR of large DNA inserts (up to 230 kb) into plant and fungal genomes.


Item Type:Article
Related URLs:
URLURL TypeDescription
http://dx.doi.org/10.1093/nar/28.7.e19DOIUNSPECIFIED
http://nar.oxfordjournals.org/content/28/7/e19.abstractPublisherUNSPECIFIED
Additional Information:© 2000 Oxford University Press. Received February 17, 1999; Revised October 8, 1999; Accepted November 10, 1999. We thank Dr Melvin Simon for critical reading of the manuscript and helpful discussions, Dr Sam Reddy for the gift of plasmid pBACKAN and Drs Eric Fang and Ralph Dean for the gift of plasmid pGGB1001.
Issue or Number:7
Record Number:CaltechAUTHORS:20111123-113725128
Persistent URL:https://resolver.caltech.edu/CaltechAUTHORS:20111123-113725128
Official Citation: Sangdun Choi, Dilara Begum, Heather Koshinsky, David W. Ow, and Rod A. Wing A new approach for the identification and cloning of genes: the pBACwich system using Cre/lox site-specific recombination Nucl. Acids Res. (2000) 28(7): e19 doi:10.1093/nar/28.7.e19
Usage Policy:No commercial reproduction, distribution, display or performance rights in this work are provided.
ID Code:27951
Collection:CaltechAUTHORS
Deposited By: Ruth Sustaita
Deposited On:23 Nov 2011 21:20
Last Modified:03 Oct 2019 03:27

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