Evaluation of the uniformity and stability of T-DNA integration and gene expression in transgenic apple plants

The generation of transgenic apple plants relies on the molecular analysis of transgene integration and expression based on polymerase chain reaction (PCR) analysis, blotting techniques and enzymatic assays on vitro leaves of putative transgenic regenerates. In order to assess the uniformity and the...

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Autores principales:	Flachowsky,Henryk, Riedel,Marko, Reim,Stefanie, Hanke,Magda-Viola
Lenguaje:	English
Publicado:	Pontificia Universidad Católica de Valparaíso 2008
Materias:	Agrobacterium tumefaciens apple chimeric tissue gene silencing Malus domestica T-DNA leakage
Acceso en línea:	http://www.scielo.cl/scielo.php?script=sci_arttext&pid=S0717-34582008000100003
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spelling	oai:scielo:S0717-345820080001000032008-10-28Evaluation of the uniformity and stability of T-DNA integration and gene expression in transgenic apple plantsFlachowsky,HenrykRiedel,MarkoReim,StefanieHanke,Magda-Viola Agrobacterium tumefaciens apple chimeric tissue gene silencing Malus domestica T-DNA leakage The generation of transgenic apple plants relies on the molecular analysis of transgene integration and expression based on polymerase chain reaction (PCR) analysis, blotting techniques and enzymatic assays on vitro leaves of putative transgenic regenerates. In order to assess the uniformity and the stability of transfer DNA (T-DNA) integration and gene expression, we studied 26 transgenic apple lines carrying the attacin E gene from Hyalophora cecropia, the β-glucuronidase gene, and the nptII gene. Plants were evaluated using standard molecular techniques, such as PCR, Southern blot, reverse transcription PCR (RT-PCR) and Enzyme Linked Immunosorbent Assay (ELISA), and propagated in vitro on non-selective antibiotic-free media for four years to mimic natural conditions in the field. In some T-lines transgene integration and expression did not remain stable; differences were also found between distinct plants of a single T-line. Individual plants with partially or completely silenced transgenes were identified as well as plants with non-detectable T-DNA. Several lines appeared chimeric or partially silenced. Although most molecular techniques can reliably detect the presence of transgenic cells, they often fail to detect mixtures of transformed and non-transformed cells, or cells with silenced transgenes. This should be taken into consideration, especially in the case of vegetatively propagated trees, where non-transformed or silenced plant parts could mistakenly be used as propagation material.info:eu-repo/semantics/openAccessPontificia Universidad Católica de ValparaísoElectronic Journal of Biotechnology v.11 n.1 20082008-01-01text/htmlhttp://www.scielo.cl/scielo.php?script=sci_arttext&pid=S0717-34582008000100003en10.4067/S0717-34582008000100003
institution	Scielo Chile
collection	Scielo Chile
language	English
topic	Agrobacterium tumefaciens apple chimeric tissue gene silencing Malus domestica T-DNA leakage
spellingShingle	Agrobacterium tumefaciens apple chimeric tissue gene silencing Malus domestica T-DNA leakage Flachowsky,Henryk Riedel,Marko Reim,Stefanie Hanke,Magda-Viola Evaluation of the uniformity and stability of T-DNA integration and gene expression in transgenic apple plants
description	The generation of transgenic apple plants relies on the molecular analysis of transgene integration and expression based on polymerase chain reaction (PCR) analysis, blotting techniques and enzymatic assays on vitro leaves of putative transgenic regenerates. In order to assess the uniformity and the stability of transfer DNA (T-DNA) integration and gene expression, we studied 26 transgenic apple lines carrying the attacin E gene from Hyalophora cecropia, the β-glucuronidase gene, and the nptII gene. Plants were evaluated using standard molecular techniques, such as PCR, Southern blot, reverse transcription PCR (RT-PCR) and Enzyme Linked Immunosorbent Assay (ELISA), and propagated in vitro on non-selective antibiotic-free media for four years to mimic natural conditions in the field. In some T-lines transgene integration and expression did not remain stable; differences were also found between distinct plants of a single T-line. Individual plants with partially or completely silenced transgenes were identified as well as plants with non-detectable T-DNA. Several lines appeared chimeric or partially silenced. Although most molecular techniques can reliably detect the presence of transgenic cells, they often fail to detect mixtures of transformed and non-transformed cells, or cells with silenced transgenes. This should be taken into consideration, especially in the case of vegetatively propagated trees, where non-transformed or silenced plant parts could mistakenly be used as propagation material.
author	Flachowsky,Henryk Riedel,Marko Reim,Stefanie Hanke,Magda-Viola
author_facet	Flachowsky,Henryk Riedel,Marko Reim,Stefanie Hanke,Magda-Viola
author_sort	Flachowsky,Henryk
title	Evaluation of the uniformity and stability of T-DNA integration and gene expression in transgenic apple plants
title_short	Evaluation of the uniformity and stability of T-DNA integration and gene expression in transgenic apple plants
title_full	Evaluation of the uniformity and stability of T-DNA integration and gene expression in transgenic apple plants
title_fullStr	Evaluation of the uniformity and stability of T-DNA integration and gene expression in transgenic apple plants
title_full_unstemmed	Evaluation of the uniformity and stability of T-DNA integration and gene expression in transgenic apple plants
title_sort	evaluation of the uniformity and stability of t-dna integration and gene expression in transgenic apple plants
publisher	Pontificia Universidad Católica de Valparaíso
publishDate	2008
url	http://www.scielo.cl/scielo.php?script=sci_arttext&pid=S0717-34582008000100003
work_keys_str_mv	AT flachowskyhenryk evaluationoftheuniformityandstabilityoftdnaintegrationandgeneexpressionintransgenicappleplants AT riedelmarko evaluationoftheuniformityandstabilityoftdnaintegrationandgeneexpressionintransgenicappleplants AT reimstefanie evaluationoftheuniformityandstabilityoftdnaintegrationandgeneexpressionintransgenicappleplants AT hankemagdaviola evaluationoftheuniformityandstabilityoftdnaintegrationandgeneexpressionintransgenicappleplants
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Evaluation of the uniformity and stability of T-DNA integration and gene expression in transgenic apple plants

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