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PhiC31 recombination system demonstrates heritable germinal transmission of site-specific excision from the Arabidopsis genome

BACKGROUND: The large serine recombinase phiC31 from broad host range Streptomyces temperate phage, catalyzes the site-specific recombination of two recognition sites that differ in sequence, typically known as attachment sites attB and attP. Previously, we characterized the phiC31 catalytic activit...

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Autores principales: Thomson, James G, Chan, Ronald, Thilmony, Roger, Yau, Yuan-Yeu, Ow, David W
Formato: Texto
Lenguaje:English
Publicado: BioMed Central 2010
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2837860/
https://www.ncbi.nlm.nih.gov/pubmed/20178628
http://dx.doi.org/10.1186/1472-6750-10-17
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author Thomson, James G
Chan, Ronald
Thilmony, Roger
Yau, Yuan-Yeu
Ow, David W
author_facet Thomson, James G
Chan, Ronald
Thilmony, Roger
Yau, Yuan-Yeu
Ow, David W
author_sort Thomson, James G
collection PubMed
description BACKGROUND: The large serine recombinase phiC31 from broad host range Streptomyces temperate phage, catalyzes the site-specific recombination of two recognition sites that differ in sequence, typically known as attachment sites attB and attP. Previously, we characterized the phiC31 catalytic activity and modes of action in the fission yeast Schizosaccharomyces pombe. RESULTS: In this work, the phiC31 recombinase gene was placed under the control of the Arabidopsis OXS3 promoter and introduced into Arabidopsis harboring a chromosomally integrated attB and attP-flanked target sequence. The phiC31 recombinase excised the attB and attP-flanked DNA, and the excision event was detected in subsequent generations in the absence of the phiC31 gene, indicating germinal transmission was possible. We further verified that the genomic excision was conservative and that introduction of a functional recombinase can be achieved through secondary transformation as well as manual crossing. CONCLUSION: The phiC31 system performs site-specific recombination in germinal tissue, a prerequisite for generating stable lines with unwanted DNA removed. The precise site-specific deletion by phiC31 in planta demonstrates that the recombinase can be used to remove selectable markers or other introduced transgenes that are no longer desired and therefore can be a useful tool for genome engineering in plants.
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spelling pubmed-28378602010-03-14 PhiC31 recombination system demonstrates heritable germinal transmission of site-specific excision from the Arabidopsis genome Thomson, James G Chan, Ronald Thilmony, Roger Yau, Yuan-Yeu Ow, David W BMC Biotechnol Research article BACKGROUND: The large serine recombinase phiC31 from broad host range Streptomyces temperate phage, catalyzes the site-specific recombination of two recognition sites that differ in sequence, typically known as attachment sites attB and attP. Previously, we characterized the phiC31 catalytic activity and modes of action in the fission yeast Schizosaccharomyces pombe. RESULTS: In this work, the phiC31 recombinase gene was placed under the control of the Arabidopsis OXS3 promoter and introduced into Arabidopsis harboring a chromosomally integrated attB and attP-flanked target sequence. The phiC31 recombinase excised the attB and attP-flanked DNA, and the excision event was detected in subsequent generations in the absence of the phiC31 gene, indicating germinal transmission was possible. We further verified that the genomic excision was conservative and that introduction of a functional recombinase can be achieved through secondary transformation as well as manual crossing. CONCLUSION: The phiC31 system performs site-specific recombination in germinal tissue, a prerequisite for generating stable lines with unwanted DNA removed. The precise site-specific deletion by phiC31 in planta demonstrates that the recombinase can be used to remove selectable markers or other introduced transgenes that are no longer desired and therefore can be a useful tool for genome engineering in plants. BioMed Central 2010-02-23 /pmc/articles/PMC2837860/ /pubmed/20178628 http://dx.doi.org/10.1186/1472-6750-10-17 Text en Copyright ©2010 Thomson et al; licensee BioMed Central Ltd. http://creativecommons.org/licenses/by/2.0 This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.
spellingShingle Research article
Thomson, James G
Chan, Ronald
Thilmony, Roger
Yau, Yuan-Yeu
Ow, David W
PhiC31 recombination system demonstrates heritable germinal transmission of site-specific excision from the Arabidopsis genome
title PhiC31 recombination system demonstrates heritable germinal transmission of site-specific excision from the Arabidopsis genome
title_full PhiC31 recombination system demonstrates heritable germinal transmission of site-specific excision from the Arabidopsis genome
title_fullStr PhiC31 recombination system demonstrates heritable germinal transmission of site-specific excision from the Arabidopsis genome
title_full_unstemmed PhiC31 recombination system demonstrates heritable germinal transmission of site-specific excision from the Arabidopsis genome
title_short PhiC31 recombination system demonstrates heritable germinal transmission of site-specific excision from the Arabidopsis genome
title_sort phic31 recombination system demonstrates heritable germinal transmission of site-specific excision from the arabidopsis genome
topic Research article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2837860/
https://www.ncbi.nlm.nih.gov/pubmed/20178628
http://dx.doi.org/10.1186/1472-6750-10-17
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