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Mol. Cells 2012; 33(1): 61-69

Published online November 29, 2011

https://doi.org/10.1007/s10059-012-2212-6

© The Korean Society for Molecular and Cellular Biology

Development of a Simple and Efficient System for Excising Selectable Markers in Arabidopsis Using a Minimal Promoter::Cre Fusion Construct

Hyun-Bi Kim1,4, Jung-Il Cho1,4, Nayeon Ryoo1, Shaohong Qu2, Guo-Liang Wang3, and Jong-Seong Jeon1,*

1Graduate School of Biotechnology and Crop Biotech Institute, Kyung Hee University, Yongin 446-701, Korea, 2Institute of Virology and Biotechnology, Zhejiang Academy of Agricultural Sciences, Hangzhou, Zhejiang 310021, China, 3Department of Plant Pathology, Ohio State University, Columbus, OH 43210, USA, 4 These authors contributed equally to this work.

Correspondence to : *Correspondence: jjeon@khu.ac.kr

Received: September 30, 2011; Revised: November 9, 2011; Accepted: November 11, 2011

Abstract

The development of rapid and efficient strategies to gen-erate selectable marker-free transgenic plants could help increase the consumer acceptance of genetically modified (GM) plants. To produce marker-free transgenic plants without conditional treatment or the genetic crossing of offspring, we have developed a rapid and convenient DNA excision method mediated by the Cre/loxP recombination system under the control of a -46 minimal CaMV 35S promoter. The results of a transient expression assay showed that -46 minimal promoter::Cre recombinase (-46::Cre) can cause the loxP-specific excision of a selectable marker, thereby connecting the 35S promoter and ?-glucuronidase (GUS) reporter gene. Analysis of stable transgenic Arabidopsis plants indicated a positive correlation between loxP-specific DNA excision and GUS expression. PCR and DNA gel-blot analysis further revealed that nine of the 10 tested T1 transgenic lines carried both excised and non-excised constructs in their genomes. In the subsequent T2 generation plants, over 30% of the individuals for each line were marker-free plants harboring the excised construct only. These results demonstrate that the -46::Cre fusion construct can be efficiently and easily utilized for producing marker-free transgenic plants.

Keywords -46 promoter, Arabidopsis, Cre, loxP, marker-free plant

Article

Research Article

Mol. Cells 2012; 33(1): 61-69

Published online January 31, 2012 https://doi.org/10.1007/s10059-012-2212-6

Copyright © The Korean Society for Molecular and Cellular Biology.

Development of a Simple and Efficient System for Excising Selectable Markers in Arabidopsis Using a Minimal Promoter::Cre Fusion Construct

Hyun-Bi Kim1,4, Jung-Il Cho1,4, Nayeon Ryoo1, Shaohong Qu2, Guo-Liang Wang3, and Jong-Seong Jeon1,*

1Graduate School of Biotechnology and Crop Biotech Institute, Kyung Hee University, Yongin 446-701, Korea, 2Institute of Virology and Biotechnology, Zhejiang Academy of Agricultural Sciences, Hangzhou, Zhejiang 310021, China, 3Department of Plant Pathology, Ohio State University, Columbus, OH 43210, USA, 4 These authors contributed equally to this work.

Correspondence to:*Correspondence: jjeon@khu.ac.kr

Received: September 30, 2011; Revised: November 9, 2011; Accepted: November 11, 2011

Abstract

The development of rapid and efficient strategies to gen-erate selectable marker-free transgenic plants could help increase the consumer acceptance of genetically modified (GM) plants. To produce marker-free transgenic plants without conditional treatment or the genetic crossing of offspring, we have developed a rapid and convenient DNA excision method mediated by the Cre/loxP recombination system under the control of a -46 minimal CaMV 35S promoter. The results of a transient expression assay showed that -46 minimal promoter::Cre recombinase (-46::Cre) can cause the loxP-specific excision of a selectable marker, thereby connecting the 35S promoter and ?-glucuronidase (GUS) reporter gene. Analysis of stable transgenic Arabidopsis plants indicated a positive correlation between loxP-specific DNA excision and GUS expression. PCR and DNA gel-blot analysis further revealed that nine of the 10 tested T1 transgenic lines carried both excised and non-excised constructs in their genomes. In the subsequent T2 generation plants, over 30% of the individuals for each line were marker-free plants harboring the excised construct only. These results demonstrate that the -46::Cre fusion construct can be efficiently and easily utilized for producing marker-free transgenic plants.

Keywords: -46 promoter, Arabidopsis, Cre, loxP, marker-free plant

Mol. Cells
May 31, 2023 Vol.46 No.5, pp. 259~328
COVER PICTURE
The alpha-helices in the lamin filaments are depicted as coils, with different subdomains distinguished by various colors. Coil 1a is represented by magenta, coil 1b by yellow, L2 by green, coil 2a by white, coil 2b by brown, stutter by cyan, coil 2c by dark blue, and the lamin Ig-like domain by grey. In the background, cells are displayed, with the cytosol depicted in green and the nucleus in blue (Ahn et al., pp. 309-318).

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