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Construction of Marker-Free Genetically Modified Maize Using a Heat-Inducible Auto-Excision Vector
Gene modification is a promising tool for plant breeding, and gradual application from the laboratory to the field. Selectable marker genes (SMG) are required in the transformation process to simplify the identification of transgenic plants; however, it is more desirable to obtain transgenic plants...
Autores principales: | , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
MDPI
2019
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6562874/ https://www.ncbi.nlm.nih.gov/pubmed/31108922 http://dx.doi.org/10.3390/genes10050374 |
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author | Du, Dengxiang Jin, Ruchang Guo, Jinjie Zhang, Fangdong |
author_facet | Du, Dengxiang Jin, Ruchang Guo, Jinjie Zhang, Fangdong |
author_sort | Du, Dengxiang |
collection | PubMed |
description | Gene modification is a promising tool for plant breeding, and gradual application from the laboratory to the field. Selectable marker genes (SMG) are required in the transformation process to simplify the identification of transgenic plants; however, it is more desirable to obtain transgenic plants without selection markers. Transgene integration mediated by site-specific recombination (SSR) systems into the dedicated genomic sites has been demonstrated in a few different plant species. Here, we present an auto-elimination vector system that uses a heat-inducible Cre to eliminate the selectable marker from transgenic maize, without the need for repeated transformation or sexual crossing. The vector combines an inducible site-specific recombinase (hsp70::Cre) that allows for the precise elimination of the selectable marker gene egfp upon heating. This marker gene is used for the initial positive selection of transgenic tissue. The egfp also functions as a visual marker to demonstrate the effectiveness of the heat-inducible Cre. A second marker gene for anthocyanin pigmentation (Rsc) is located outside of the region eliminated by Cre and is used for the identification of transgenic offspring in future generations. Using the heat-inducible auto-excision vector, marker-free transgenic maize plants were obtained in a precisely controlled genetic modification process. Genetic and molecular analyses indicated that the inducible auto-excision system was tightly controlled, with highly efficient DNA excision, and provided a highly reliable method to generate marker-free transgenic maize. |
format | Online Article Text |
id | pubmed-6562874 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2019 |
publisher | MDPI |
record_format | MEDLINE/PubMed |
spelling | pubmed-65628742019-06-17 Construction of Marker-Free Genetically Modified Maize Using a Heat-Inducible Auto-Excision Vector Du, Dengxiang Jin, Ruchang Guo, Jinjie Zhang, Fangdong Genes (Basel) Article Gene modification is a promising tool for plant breeding, and gradual application from the laboratory to the field. Selectable marker genes (SMG) are required in the transformation process to simplify the identification of transgenic plants; however, it is more desirable to obtain transgenic plants without selection markers. Transgene integration mediated by site-specific recombination (SSR) systems into the dedicated genomic sites has been demonstrated in a few different plant species. Here, we present an auto-elimination vector system that uses a heat-inducible Cre to eliminate the selectable marker from transgenic maize, without the need for repeated transformation or sexual crossing. The vector combines an inducible site-specific recombinase (hsp70::Cre) that allows for the precise elimination of the selectable marker gene egfp upon heating. This marker gene is used for the initial positive selection of transgenic tissue. The egfp also functions as a visual marker to demonstrate the effectiveness of the heat-inducible Cre. A second marker gene for anthocyanin pigmentation (Rsc) is located outside of the region eliminated by Cre and is used for the identification of transgenic offspring in future generations. Using the heat-inducible auto-excision vector, marker-free transgenic maize plants were obtained in a precisely controlled genetic modification process. Genetic and molecular analyses indicated that the inducible auto-excision system was tightly controlled, with highly efficient DNA excision, and provided a highly reliable method to generate marker-free transgenic maize. MDPI 2019-05-17 /pmc/articles/PMC6562874/ /pubmed/31108922 http://dx.doi.org/10.3390/genes10050374 Text en © 2019 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (http://creativecommons.org/licenses/by/4.0/). |
spellingShingle | Article Du, Dengxiang Jin, Ruchang Guo, Jinjie Zhang, Fangdong Construction of Marker-Free Genetically Modified Maize Using a Heat-Inducible Auto-Excision Vector |
title | Construction of Marker-Free Genetically Modified Maize Using a Heat-Inducible Auto-Excision Vector |
title_full | Construction of Marker-Free Genetically Modified Maize Using a Heat-Inducible Auto-Excision Vector |
title_fullStr | Construction of Marker-Free Genetically Modified Maize Using a Heat-Inducible Auto-Excision Vector |
title_full_unstemmed | Construction of Marker-Free Genetically Modified Maize Using a Heat-Inducible Auto-Excision Vector |
title_short | Construction of Marker-Free Genetically Modified Maize Using a Heat-Inducible Auto-Excision Vector |
title_sort | construction of marker-free genetically modified maize using a heat-inducible auto-excision vector |
topic | Article |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6562874/ https://www.ncbi.nlm.nih.gov/pubmed/31108922 http://dx.doi.org/10.3390/genes10050374 |
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