Contributions of in situ hybridization of sRNA to the study on spatio-temporal gene expression in Hevea brasiliensis
Since the dawn of the genomics era, much research has focused on functional studies of genes of interest. In situ hybridization is a method that can be used to precisely localize the expression of a gene in tissues and cells. This article describes how the method has been adapted to the analysis of Hevea tissues. Initially, the conventional method of digoxigenin detection with NBT/BCIP revealed the expression of strongly expressed genes in tissues of different differentiation intensity. A new digoxigenin detection method using Alexa488 fluorochrome-labelled antibodies has been used to detect the expression of more weakly expressed genes. This method, combined with observation under a confocal microscope, has enabled very precise localization of expression. Some examples of in situ hybridization use are described for Hevea gene expression in somatic plantlets and shoot bark: the uidA gene in callus and transgenic somatic plantlets, the HEV2.1 gene encoding hevein and the ACO-H5 gene involved in ethylene metabolism. Cell imaging methods therefore open up fundamental prospects for studying the different molecular mechanisms involved in some agronomic traits of Hevea.
| Main Authors: | , , , , , , |
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| Format: | conference_item biblioteca |
| Language: | eng |
| Published: |
s.n.
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| Subjects: | F30 - Génétique et amélioration des plantes, Hevea brasiliensis, http://aims.fao.org/aos/agrovoc/c_3589, |
| Online Access: | http://agritrop.cirad.fr/548938/ http://agritrop.cirad.fr/548938/1/document_548938.pdf |
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| Summary: | Since the dawn of the genomics era, much research has focused on functional studies of genes of interest. In situ hybridization is a method that can be used to precisely localize the expression of a gene in tissues and cells. This article describes how the method has been adapted to the analysis of Hevea tissues. Initially, the conventional method of digoxigenin detection with NBT/BCIP revealed the expression of strongly expressed genes in tissues of different differentiation intensity. A new digoxigenin detection method using Alexa488 fluorochrome-labelled antibodies has been used to detect the expression of more weakly expressed genes. This method, combined with observation under a confocal microscope, has enabled very precise localization of expression. Some examples of in situ hybridization use are described for Hevea gene expression in somatic plantlets and shoot bark: the uidA gene in callus and transgenic somatic plantlets, the HEV2.1 gene encoding hevein and the ACO-H5 gene involved in ethylene metabolism. Cell imaging methods therefore open up fundamental prospects for studying the different molecular mechanisms involved in some agronomic traits of Hevea. |
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