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Identification of candidate genes for drought tolerance in coffee by high-throughput sequencing in the shoot apex of different Coffea arabica cultivars.

Mofatto LS, Carneiro Fde A, Vieira NG, Duarte KE, Vidal RO, Alekcevetch JC, Cotta MG, Verdeil JL, Lapeyre-Montes F, Lartaud M, Leroy T, De Bellis F, Pot D, Rodrigues GC, Carazzolle MF, Pereira GA, Andrade AC, Marraccini P - BMC Plant Biol. (2016)

Bottom Line: Genetic diversity for drought tolerance exists within the coffee genus.This may have been related to the thicker cuticle observed on the abaxial leaf surface in IAPAR59 compared to Rubi.The identification of these genes should help advance our understanding of the genetic determinism of drought tolerance in coffee.

View Article: PubMed Central - PubMed

Affiliation: Laboratório de Genômica e Expressão (LGE), Departamento de Genética e Evolução, Instituto de Biologia/UNICAMP, Cidade Universitária Zeferino Vaz, 13083-970, Campinas, SP, Brazil.

ABSTRACT

Background: Drought is a widespread limiting factor in coffee plants. It affects plant development, fruit production, bean development and consequently beverage quality. Genetic diversity for drought tolerance exists within the coffee genus. However, the molecular mechanisms underlying the adaptation of coffee plants to drought are largely unknown. In this study, we compared the molecular responses to drought in two commercial cultivars (IAPAR59, drought-tolerant and Rubi, drought-susceptible) of Coffea arabica grown in the field under control (irrigation) and drought conditions using the pyrosequencing of RNA extracted from shoot apices and analysing the expression of 38 candidate genes.

Results: Pyrosequencing from shoot apices generated a total of 34.7 Mbp and 535,544 reads enabling the identification of 43,087 clusters (41,512 contigs and 1,575 singletons). These data included 17,719 clusters (16,238 contigs and 1,575 singletons) exclusively from 454 sequencing reads, along with 25,368 hybrid clusters assembled with 454 sequences. The comparison of DNA libraries identified new candidate genes (n = 20) presenting differential expression between IAPAR59 and Rubi and/or drought conditions. Their expression was monitored in plagiotropic buds, together with those of other (n = 18) candidates genes. Under drought conditions, up-regulated expression was observed in IAPAR59 but not in Rubi for CaSTK1 (protein kinase), CaSAMT1 (SAM-dependent methyltransferase), CaSLP1 (plant development) and CaMAS1 (ABA biosynthesis). Interestingly, the expression of lipid-transfer protein (nsLTP) genes was also highly up-regulated under drought conditions in IAPAR59. This may have been related to the thicker cuticle observed on the abaxial leaf surface in IAPAR59 compared to Rubi.

Conclusions: The full transcriptome assembly of C. arabica, followed by functional annotation, enabled us to identify differentially expressed genes related to drought conditions. Using these data, candidate genes were selected and their differential expression profiles were confirmed by qPCR experiments in plagiotropic buds of IAPAR59 and Rubi under drought conditions. As regards the genes up-regulated under drought conditions, specifically in the drought-tolerant IAPAR59, several corresponded to orphan genes but also to genes coding proteins involved in signal transduction pathways, as well as ABA and lipid metabolism, for example. The identification of these genes should help advance our understanding of the genetic determinism of drought tolerance in coffee.

No MeSH data available.


Related in: MedlinePlus

Expression of nsLTP genes. Expression of CaLTP1-CaLTP2 (CaCe: white isobars), CaLTP3 (CaCc: grey isobars) and all (CaLTP1, CaLTP2 and CaLTP3: black isobars) genes was analysed by qPCR in plagiotropic buds of Rubi (RUB) and IAPAR59 (I59) cultivars of C. arabica grown under control (C) and drought (D) conditions, using the LTP-FT/LTP-R2, LTP-FT/LTP-R1 and LTP-F100/LTP-R100 primer pairs, respectively [37]. Expression levels are expressed in arbitrary units (AU) of nsLTP genes using the expression of the CaUBQ10 gene as the endogenous control and RUB-C (with LTP100 primers) as the reference sample (Relative expression = 1). Values of three technical replications are presented as mean ± SD (bar)
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Fig5: Expression of nsLTP genes. Expression of CaLTP1-CaLTP2 (CaCe: white isobars), CaLTP3 (CaCc: grey isobars) and all (CaLTP1, CaLTP2 and CaLTP3: black isobars) genes was analysed by qPCR in plagiotropic buds of Rubi (RUB) and IAPAR59 (I59) cultivars of C. arabica grown under control (C) and drought (D) conditions, using the LTP-FT/LTP-R2, LTP-FT/LTP-R1 and LTP-F100/LTP-R100 primer pairs, respectively [37]. Expression levels are expressed in arbitrary units (AU) of nsLTP genes using the expression of the CaUBQ10 gene as the endogenous control and RUB-C (with LTP100 primers) as the reference sample (Relative expression = 1). Values of three technical replications are presented as mean ± SD (bar)

Mentions: The expression of Type II nsLTP-encoding genes was also monitored using the primer pairs LTP-FT/LTP-R1 (specific to the CaLTP1 and CaLTP2 genes from the C. eugenioides sub-genome of C. arabica, hereafter referred to as CaCe), LTP-FT/LTP-R2 (specific to CaLTP3 genes from the C. canephora of C. arabica, hereafter CaCc) and LTP-F100/LTP-R100 recognizing all homologous genes [32]. No expression of nsLTP genes was detected under the control conditions in both cultivars (Fig. 5). However, expression of nsLTP genes was highly up-regulated in IAPAR59 but not in Rubi under drought conditions. It is worth noting that the CaLTP1-CaLTP2 and CaLTP3 genes were co-expressed in IAPAR59, and that the expression of CaCc genes was slightly higher than that of CaCe genes.Fig. 5


Identification of candidate genes for drought tolerance in coffee by high-throughput sequencing in the shoot apex of different Coffea arabica cultivars.

Mofatto LS, Carneiro Fde A, Vieira NG, Duarte KE, Vidal RO, Alekcevetch JC, Cotta MG, Verdeil JL, Lapeyre-Montes F, Lartaud M, Leroy T, De Bellis F, Pot D, Rodrigues GC, Carazzolle MF, Pereira GA, Andrade AC, Marraccini P - BMC Plant Biol. (2016)

Expression of nsLTP genes. Expression of CaLTP1-CaLTP2 (CaCe: white isobars), CaLTP3 (CaCc: grey isobars) and all (CaLTP1, CaLTP2 and CaLTP3: black isobars) genes was analysed by qPCR in plagiotropic buds of Rubi (RUB) and IAPAR59 (I59) cultivars of C. arabica grown under control (C) and drought (D) conditions, using the LTP-FT/LTP-R2, LTP-FT/LTP-R1 and LTP-F100/LTP-R100 primer pairs, respectively [37]. Expression levels are expressed in arbitrary units (AU) of nsLTP genes using the expression of the CaUBQ10 gene as the endogenous control and RUB-C (with LTP100 primers) as the reference sample (Relative expression = 1). Values of three technical replications are presented as mean ± SD (bar)
© Copyright Policy - OpenAccess
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC4837521&req=5

Fig5: Expression of nsLTP genes. Expression of CaLTP1-CaLTP2 (CaCe: white isobars), CaLTP3 (CaCc: grey isobars) and all (CaLTP1, CaLTP2 and CaLTP3: black isobars) genes was analysed by qPCR in plagiotropic buds of Rubi (RUB) and IAPAR59 (I59) cultivars of C. arabica grown under control (C) and drought (D) conditions, using the LTP-FT/LTP-R2, LTP-FT/LTP-R1 and LTP-F100/LTP-R100 primer pairs, respectively [37]. Expression levels are expressed in arbitrary units (AU) of nsLTP genes using the expression of the CaUBQ10 gene as the endogenous control and RUB-C (with LTP100 primers) as the reference sample (Relative expression = 1). Values of three technical replications are presented as mean ± SD (bar)
Mentions: The expression of Type II nsLTP-encoding genes was also monitored using the primer pairs LTP-FT/LTP-R1 (specific to the CaLTP1 and CaLTP2 genes from the C. eugenioides sub-genome of C. arabica, hereafter referred to as CaCe), LTP-FT/LTP-R2 (specific to CaLTP3 genes from the C. canephora of C. arabica, hereafter CaCc) and LTP-F100/LTP-R100 recognizing all homologous genes [32]. No expression of nsLTP genes was detected under the control conditions in both cultivars (Fig. 5). However, expression of nsLTP genes was highly up-regulated in IAPAR59 but not in Rubi under drought conditions. It is worth noting that the CaLTP1-CaLTP2 and CaLTP3 genes were co-expressed in IAPAR59, and that the expression of CaCc genes was slightly higher than that of CaCe genes.Fig. 5

Bottom Line: Genetic diversity for drought tolerance exists within the coffee genus.This may have been related to the thicker cuticle observed on the abaxial leaf surface in IAPAR59 compared to Rubi.The identification of these genes should help advance our understanding of the genetic determinism of drought tolerance in coffee.

View Article: PubMed Central - PubMed

Affiliation: Laboratório de Genômica e Expressão (LGE), Departamento de Genética e Evolução, Instituto de Biologia/UNICAMP, Cidade Universitária Zeferino Vaz, 13083-970, Campinas, SP, Brazil.

ABSTRACT

Background: Drought is a widespread limiting factor in coffee plants. It affects plant development, fruit production, bean development and consequently beverage quality. Genetic diversity for drought tolerance exists within the coffee genus. However, the molecular mechanisms underlying the adaptation of coffee plants to drought are largely unknown. In this study, we compared the molecular responses to drought in two commercial cultivars (IAPAR59, drought-tolerant and Rubi, drought-susceptible) of Coffea arabica grown in the field under control (irrigation) and drought conditions using the pyrosequencing of RNA extracted from shoot apices and analysing the expression of 38 candidate genes.

Results: Pyrosequencing from shoot apices generated a total of 34.7 Mbp and 535,544 reads enabling the identification of 43,087 clusters (41,512 contigs and 1,575 singletons). These data included 17,719 clusters (16,238 contigs and 1,575 singletons) exclusively from 454 sequencing reads, along with 25,368 hybrid clusters assembled with 454 sequences. The comparison of DNA libraries identified new candidate genes (n = 20) presenting differential expression between IAPAR59 and Rubi and/or drought conditions. Their expression was monitored in plagiotropic buds, together with those of other (n = 18) candidates genes. Under drought conditions, up-regulated expression was observed in IAPAR59 but not in Rubi for CaSTK1 (protein kinase), CaSAMT1 (SAM-dependent methyltransferase), CaSLP1 (plant development) and CaMAS1 (ABA biosynthesis). Interestingly, the expression of lipid-transfer protein (nsLTP) genes was also highly up-regulated under drought conditions in IAPAR59. This may have been related to the thicker cuticle observed on the abaxial leaf surface in IAPAR59 compared to Rubi.

Conclusions: The full transcriptome assembly of C. arabica, followed by functional annotation, enabled us to identify differentially expressed genes related to drought conditions. Using these data, candidate genes were selected and their differential expression profiles were confirmed by qPCR experiments in plagiotropic buds of IAPAR59 and Rubi under drought conditions. As regards the genes up-regulated under drought conditions, specifically in the drought-tolerant IAPAR59, several corresponded to orphan genes but also to genes coding proteins involved in signal transduction pathways, as well as ABA and lipid metabolism, for example. The identification of these genes should help advance our understanding of the genetic determinism of drought tolerance in coffee.

No MeSH data available.


Related in: MedlinePlus