Agrobacterium based stable transformation procedures are well established in the plant science community. In particular the floral dip method is a commonly used transformation protocol applied to the model plant, Arabidopsis thaliana. Nevertheless, often the need exists for a simpler analysis of gene-GFP fusions as control of the constructs that will be used in the creation of transgenic lines or to just obtain data from them. Analysis of GFP-fusions has become an essential step in the characterization of gene functional aspects related to tissue, cellular localization and associated protein movement. The available methodologies though are often impaired by complex protocols and technical pitfalls. Thus, a procedure based on a common and widely used system of in vitro Arabidopsis growth could be very useful. We needed a quick and simple method for analysis of gene-product-GFP fusions. Thus we decided to establish a simpler procedure for Agrobacteriummediated transformation of Arabidopsis. Results demonstrating the effectiveness of this method using a 35S-CaMV promoter-GFP construct and its validation for sub-cellular localization will be presented.

SIMPLE AGROBACTERIUM-BASED TRANSFORMATION PROCEDURE USEFUL FOR GFP-FUSION

RODRIGUES POUSADA, RENATO ALBERTO;
2009

Abstract

Agrobacterium based stable transformation procedures are well established in the plant science community. In particular the floral dip method is a commonly used transformation protocol applied to the model plant, Arabidopsis thaliana. Nevertheless, often the need exists for a simpler analysis of gene-GFP fusions as control of the constructs that will be used in the creation of transgenic lines or to just obtain data from them. Analysis of GFP-fusions has become an essential step in the characterization of gene functional aspects related to tissue, cellular localization and associated protein movement. The available methodologies though are often impaired by complex protocols and technical pitfalls. Thus, a procedure based on a common and widely used system of in vitro Arabidopsis growth could be very useful. We needed a quick and simple method for analysis of gene-product-GFP fusions. Thus we decided to establish a simpler procedure for Agrobacteriummediated transformation of Arabidopsis. Results demonstrating the effectiveness of this method using a 35S-CaMV promoter-GFP construct and its validation for sub-cellular localization will be presented.
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Utilizza questo identificativo per citare o creare un link a questo documento: http://hdl.handle.net/11697/28061
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