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Functional analysis of CcDREB1D promoter region from haplotypes of Coffea canephora through genetic transformation of Nicotiana tabacum

Aquino Sinara O., Carneiro Fernanda A, Rego Erica C.S., Alves Gabriel Sergio Costa, Duarte Karoline Estefani, Marraccini Pierre, Andrade Alan Carvalho. 2014. Functional analysis of CcDREB1D promoter region from haplotypes of Coffea canephora through genetic transformation of Nicotiana tabacum. In : 11th Solanaceae Conference: book of abstracts, Porto Seguro, Brazil, November 2-6, 2014. Embrapa, UESC, UFLA, Ciba. s.l. : s.n., Résumé, p. 168. Solanaceae Conference. 11, Porto Seguro, Brésil, 2 November 2014/6 November 2014.

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Abstract : Recent studies in coffee resulted in the identification of many candidate genes for drought tolerance characterized by differential expression profiles in leaves of drought-tolerant and susceptible clones of Coffea canephora Conilon. Among those are found the genes involved in the water stress response pathway, such as CcDREB1D (coding for DREB transcription factors) that showed higher expression under water stress conditions in the leaves of clone 14 (droughttolerant) than in those of the clone 22 (drought-sensitive). After sequencing, nucleic polymorphisms were identified in the promoter regions of CcDREB1D gene of clones 14 and 22, indicating the presence of three haplotypes (15, 16 and 17) of this promoter. With the aim of studying the participation of these polymorphisms in the response to drought, several genetic constructions of the CcDREB1D promoter regions were made in the pBI101 binary vector and tested via genetic transformation of Nicotiana tabacum cv. SRI, by evaluating the capacity of such fragments in controlling the expression of the ?-glucoronidase (uidA) reporter gene. For the constructions containing the longest versions (D) of the pDREB1D, a basal expression of uidA gene was observed in both leaves and roots of T0 plants grown without drought stress. To see if CcDREB1D haplotypes would respond to abiotic stresses, T0 tobacco plants were submitted to dehydration and elevated temperature assays, and subsequently analyzed for the expression of the uidA reporter gene by GUS histochemical tests and real-time quantitative PCR (RT-qPCR). A slight induction of the uidA gene was confirmed in the leaves of T0 plants transformed with pD22-hp17D. However, gene expression levels were much lower than those measured in plants transformed with the positive control (pBI121). No induction of the reporter gene was observed in plants transformed with the different constructions containing the other haplotypes (15 and 16) of the CcDREB1D promoter. Altogether, these results showed that (i) the pDREB1D promoters of C. canephora are weak in tobacco and that (ii) the haplotype 17 of this promoter, derived from C. canephora clone 22, was induced with abiotic stresses in the tobacco leaves. This indicates that the molecular mechanisms implicated in the regulation of the gene expression in response to drought are (at least partially) conserved between coffee and tobacco plants and that the functioning of the pDREB1D promoters from coffee clones 14 and 22 is different in tobacco, suggesting that the polymorphisms previously identified are important in regulating these promoters. (Texte intégral)

Classification Agris : F30 - Plant genetics and breeding
H50 - Miscellaneous plant disorders
F60 - Plant physiology and biochemistry

Auteurs et affiliations

  • Aquino Sinara O., EMBRAPA (BRA)
  • Carneiro Fernanda A, EMBRAPA (BRA)
  • Rego Erica C.S., EMBRAPA (BRA)
  • Alves Gabriel Sergio Costa, UFLA (BRA)
  • Duarte Karoline Estefani, EMBRAPA (BRA)
  • Marraccini Pierre, CIRAD-BIOS-UMR AGAP (BRA) ORCID: 0000-0001-7637-6811
  • Andrade Alan Carvalho, EMBRAPA (BRA)

Source : Cirad - Agritrop (https://agritrop.cirad.fr/574951/)

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