Genetics and Molecular Biology

URI permanente para esta coleçãohttps://thoth.dti.ufv.br/handle/123456789/13110

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Caffeine content of Ethiopian Coffea arabica beans
(Sociedade Brasileira de Genética, 2000) Silvarolla, Maria Bernadete; Mazzafera, Paulo; Lima, Marinez Muraro Alves de
The coffee germplasm bank of the Instituto Agronômico de Campinas has many Coffea arabica accessions from Ethiopia, which is considered the primary center of genetic diversity in coffee plants. An evaluation of the caffeine content of beans from 99 progenies revealed intra- and inter-progeny variability. In 68 progenies from the Kaffa region we found caffeine values in the range 0.46-2.82% (mean 1.18%), and in 22 progenies from Illubabor region these values ranged from 0.42 to 2.90% (mean 1.10%). This variability could be exploited in a breeding program aimed at producing beans with low-caffeine content.
Genetic polymorphism among 14 elite Coffea arabica L. cultivars using RAPD markers associated with restriction digestion
(Sociedade Brasileira de Genética, 2003) Sera, Tumoru; Ruas, Paulo Maurício; Ruas, Claudete de Fátima; Diniz, Leandro Eugênio Cardamone; Carvalho, Valdemar de Paula; Rampim, Leandro; Ruas, Eduardo Augusto; Silveira, Sheila Recepute da
Knowledge of the genetic variability among genotypes is important for the transfer of useful genes and to maximize the use of available germplasm resources. This study was carried out to assess the genetic variability of 14 elite Coffea arabica cultivars using random amplified polymorphic DNA (RAPD) associated with a prior digestion of genomic DNA with restriction endonucleases. The accessions were obtained from the Coffea collection maintained at the Instituto Agronômico do Paraná (IAPAR), located in Londrina, Paraná, Brazil. Twenty-four informative RAPD primers, used in association with restriction enzymes, yielded 330 reproducible and scorable DNA bands, of which 224 (68%) were polymorphic. The amplified products were used to estimate the genetic variability using Dice’s similarity coefficient. The data matrix was converted to a dendrogram and a three-dimensional plot using principal coordinate analysis. The accessions studied were separated into clusters in a manner that was consistent with the known pedigree. The associations obtained in the dendrogram and in the principal coordinate analysis plot suggest the probable origin of the Kattimor cultivar. The RAPD technique associated with restriction digestion was proved to be a useful tool for genetic characterization of C. arabica genotypes making an important contribution to the application of molecular markers to coffee breeding.
Assessment of genetic variability within and among coffee progenies and cultivars using RAPD markers
(Sociedade Brasileira de Genética, 2003) Silveira, Sheila Recepute; Ruas, Paulo Maurício; Ruas, Claudete de Fátima; Sera, Tumoru; Carvalho, Valdemar de Paula; Coelho, Alexandre Siqueira Guedes
The RAPD technique associated with restriction digestion of genomic DNA was used to assess the genetic variability within and among nine populations of Coffea arabica, including six progenies belonging to the Sarchimor germplasm, the progeny PR 77054-40-10 (Catuaí Vermelho IAC 81 x Icatu), and two commercial cultivars (IAPAR 59 and Catuaí Vermelho IAC-81). These populations were evaluated using analysis of molecular variance (AMOVA), genetic similarity among progenies, and percentage of polymorphic loci. A total of 99 RAPD markers were evaluated of which 67 (67.67%) were polymorphic. AMOVA showed that 38.5% and 61.5% of the genetic variation was distributed among and within populations, respectively. The fixation index (FST) of the genotypes was 0.385. The mean genetic variability estimated within populations ranged from 15.58 (IAPAR 59) to 8.27 (Catuaí Vermelho IAC 81). A distinct level of genetic variability was revealed for each of the coffee progenies and varieties studied. The methodology used in this investigation was useful to determine the genetic variability within and among C. arabica L. populations providing significant information for coffee breeding.
Genetic relationship in Coffea species and parentage determination of interspecific hybrids using ISSR (Inter- Simple Sequence Repeat) markers
(Sociedade Brasileira de Genética, 2003) Ruas, Paulo M.; Ruas, Claudete F.; Rampim, Leandro; Carvalho, Valdemar P.; Ruas, Eduardo A.; Sera, Tumoru
Inter-simple sequence repeat (ISSR) markers were used to evaluate genetic divergence among eight Coffea species and to identify the parentage of six interspecific hybrids. A total of 14 primers which contained different simple sequence repeats (SSR) were used as single primers or combined in pairs and tested for PCR amplifications. Two hundred and thirty highly reproducible fragments were amplified, which were then used to estimate the genetic similarity and to cluster the Coffea species and hybrids. High levels of interspecific genetic variation were revealed. The dinucleotide motif (GA)9T combined with other di- tri- and tetra-nucleotides produced a greater number of DNA fragments, mostly polymorphics, suggesting a high frequency of the poly GA microsatellite motifs in the Coffea genomes. The genetic similarity ranged from 0.25 between C. racemosa and C. liberica var. dewevrei to 0.86 between C. arabica var. arabica and Hybrid N. 2. The C. arabica species shared most of its markers with five of the six hybrids suggesting that it is the most likely candidate as one of the progenitors of those hybrids. These results revealed that ISSR markers could be efficiently used for genetic differentiation of the Coffea species and to identify the parentage of Coffea interspecific hybrids.
Effects of caffeine and used coffee grounds on biological features of Aedes aegypti (Diptera, Culicidae) and their possible use in alternative control
(Sociedade Brasileira de Genética, 2003) Laranja, Alessandra Theodoro; Manzatto, Antonio José; Bicudo, Hermione Elly Melara de Campos
Caffeine and used coffee grounds completely blocked the development of Aedes aegypti in the early stages, in treatments with the concentrations 1.0 mg/mL and 50 mg/mL, respectively. More advanced stages and even adults were obtained in lower concentrations of both substances, enabling observations to be made of mortality rate, longevity and esterase patterns. The experiments involved treatments using either eggs or 3rd instar larvae (L3), with or without the addition of fish food. Mortality rates prior to the adult stage and adult longevity were significantly different in the comparisons among treatments, in every kind of experiment, but in those using L3 larvae, their percentages were smaller. Observations of the time of larva and adult onset suggested that developmental time was also delayed in treatments with both substances. The addition of fish food increased significantly the number of adults produced in caffeine 0.2 and in the control, but in used coffee grounds, the opposite effect occurred. Longevity was apparently not affected by the addition of food, except again in coffee grounds, in which it decreased. In an attempt to detect a mechanism involved in the action of caffeine and coffee grounds, esterases (enzymes involved in the detoxification of xenobiotics) were analyzed in polyacrylamide gels of treated 4th instar larvae (L4). In treatments with both substances, the expression of some carboxylesterases was affected, suggesting that they may be involved in the observed impairment.
Caffeine inheritance in interspecific hybrids of Coffea arabica x Coffea canephora (Gentianales, Rubiaceae)
(Sociedade Brasileira de Genética, 2008) Priolli, Regina H.G.; Mazzafera, Paulo; Siqueira, Walter J.; Möller, Milene; Zucchi, Maria Imaculada; Ramos, Luis Carlos S.; Gallo, Paulo B.; Colombo, Carlos A.
Caffeine inheritance was investigated in F2 and BC1F1 generations between Coffea arabica var. Bourbon Vermelho (BV) and Coffea canephora var. Robusta 4x (R4x). The caffeine content of seeds and leaves was determined during 2004 and 2005. Microsatellite loci-markers were used to deduce the meiotic pattern of chromosome pairing of tetraploid interspecific hybrids. Genetic analysis indicated that caffeine content in seeds was quantitatively inherited and controlled by genes with additive effects. The estimates of broad-sense heritability of caffeine content in seeds were high for both generations. In coffee leaves, the caffeine content (BSH) from the same populations showed transgressive segregants with enhanced levels and high BSH. Segregation of loci-markers in BC1F1 populations showed that the ratios of the gametes genotype did not differ significantly from those expected assuming random associations and tetrasomic inheritance. The results confirm the existence of distinct mechanisms controlling the caffeine content in seeds and leaves, the gene exchange between the C. arabica BV and C. canephora R4x genomes and favorable conditions for improving caffeine content in this coffee population.
Altered expression of the caffeine synthase gene in a naturally caffeine-free mutant of Coffea arabica
(Sociedade Brasileira de Genética, 2009) Maluf, Mirian Perez; Silva, Carla Cristina da; Oliveira, Michelle de Paula Abreu de; Tavares, Aline Gomes; Silvarolla, Maria Bernadete; Guerreiro Filho, Oliveiro
In this work, we studied the biosynthesis of caffeine by examining the expression of genes involved in this biosynthetic pathway in coffee fruits containing normal or low levels of this substance. The amplification of gene-specific transcripts during fruit development revealed that low-caffeine fruits had a lower expression of the theobromine synthase and caffeine synthase genes and also contained an extra transcript of the caffeine synthase gene. This extra transcript contained only part of exon 1 and all of exon 3. The sequence of the mutant caffeine synthase gene revealed the substitution of isoleucine for valine in the enzyme active site that probably interfered with enzymatic activity. These findings indicate that the absence of caffeine in these mutants probably resulted from a combination of transcriptional regulation and the presence of mutations in the caffeine synthase amino acid sequence.
Development of microsatellite markers for identifying Brazilian Coffea arabica varieties
(Sociedade Brasileira de Genética, 2010) Vieira, Elisa S.N.; Pinho, Édila V. de R. Von; Carvalho, Maria G.G.; Esselink, Danny G.; Vosman, Ben
Microsatellite markers, also known as SSRs (Simple Sequence Repeats), have proved to be excellent tools for identifying variety and determining genetic relationships. A set of 127 SSR markers was used to analyze genetic similarity in twenty five Coffea arabica varieties. These were composed of nineteen commercially important Brazilians and six interspecific hybrids of Coffea arabica, Coffea canephora and Coffea liberica. The set used comprised 52 newly developed SSR markers derived from microsatellite enriched libraries, 56 designed on the basis of coffee SSR sequences available from public databases, 6 already published, and 13 universal chloroplast microsatellite markers. Only 22 were polymorphic, these detecting 2-7 alleles per marker, an average of 2.5. Based on the banding patterns generated by polymorphic SSR loci, the set of twenty-five coffee varieties were clustered into two main groups, one composed of only Brazilian varieties, and the other of interspecific hybrids, with a few Brazilians. Color mutants could not be separated. Clustering was in accordance with material genealogy thereby revealing high similarity.
In silico identification of coffee genome expressed sequences potentially associated with resistance to diseases
(Sociedade Brasileira de Genética, 2010) Alvarenga, Samuel Mazzinghy; Caixeta, Eveline Teixeira; Hufnagel, Bárbara; Thiebaut, Flávia; Maciel-Zambolim, Eunize; Zambolim, Laércio; Sakiyama, Ney Sussumu
Sequences potentially associated with coffee resistance to diseases were identified by in silico analyses using the database of the Brazilian Coffee Genome Project (BCGP). Keywords corresponding to plant resistance mechanisms to pathogens identified in the literature were used as baits for data mining. Expressed sequence tags (ESTs) related to each of these keywords were identified with tools available in the BCGP bioinformatics platform. A total of 11,300 ESTs were mined. These ESTs were clustered and formed 979 EST-contigs with similarities to chitinases, kinases, cytochrome P450 and nucleotide binding site-leucine rich repeat (NBS-LRR) proteins, as well as with proteins related to disease resistance, pathogenesis, hypersensitivity response (HR) and plant defense responses to diseases. The 140 EST-contigs identified through the keyword NBS-LRR were classified according to function. This classification allowed association of the predicted products of EST-contigs with biological processes, including host defense and apoptosis, and with molecular functions such as nucleotide binding and signal transducer activity. Fisher’s exact test was used to examine the significance of differences in contig expression between libraries representing the responses to biotic stress challenges and other libraries from the BCGP. This analysis revealed seven contigs highly similar to catalase, chitinase, protein with a BURP domain and unknown proteins. The involvement of these coffee proteins in plant responses to disease is discussed.
Identification of novel and conserved microRNAs in Coffea canephora and Coffea arabica
(Sociedade Brasileira de Genética, 2014) Loss-Morais, Guilherme; Ferreira, Daniela C.R.; Margis, Rogério; Alves-Ferreira, Márcio; Corrêa, Régis L.
As microRNAs (miRNAs) are important regulators of many biological processes, a series of small RNAomes from plants have been produced in the last decade. However, miRNA data from several groups of plants are still lacking, including some economically important crops. Here microRNAs from Coffea canephora leaves were profiled and 58 unique sequences belonging to 33 families were found, including two novel microRNAs that have never been described before in plants. Some of the microRNA sequences were also identified in Coffea arabica that, together with C. canephora, correspond to the two major sources of coffee production in the world. The targets of almost all miRNAs were also predicted on coffee expressed sequences. This is the first report of novel miRNAs in the genus Coffea, and also the first in the plant order Gentianales. The data obtained establishes the basis for the understanding of the complex miRNA-target network on those two important crops.
Galactinol synthase transcriptional profile in two genotypes of Coffea canephora with contrasting tolerance to drought
(Sociedade Brasileira de Genética, 2015) Santos, Tiago Benedito Dos; Lima, Rogério Barbosa de; Nagashima, Getúlio Takashi; Petkowicz, Carmen Lucia de Oliveira; Carpentieri-Pípolo, Valéria; Pereira, Luiz Filipe Protasio; Domingues, Douglas Silva; Vieira, Luiz Gonzaga Esteves
Increased synthesis of galactinol and raffinose family oligosaccharides (RFOs) has been reported in vegetative tissues in response to a range of abiotic stresses. In this work, we evaluated the transcriptional profile of a Coffea canephora galactinol synthase gene (CcGolS1) in two clones that differed in tolerance to water deficit in order to assess the contribution of this gene to drought tolerance. The expression of CcGolS1 in leaves was differentially regulated by water deficit, depending on the intensity of stress and the genotype. In clone 109A (drought-susceptible), the abundance of CcGolS1 transcripts decreased upon exposure to drought, reaching minimum values during recovery from severe water deficit and stress. In contrast, CcGolS1 gene expression in clone 14 (drought-tolerant) was stimulated by water deficit. Changes in galactinol and RFO content did not correlate with variation in the steady-state transcript level. However, the magnitude of increase in RFO accumulation was higher in the tolerant cultivar, mainly under severe water deficit. The finding that the drought-tolerant coffee clone showed enhanced accumulation of CcGolS1 transcripts and RFOs under water deficit suggests the possibility of using this gene to improve drought tolerance in this important crop.
Genome-wide identification, classification and transcriptional analysis of nitrate and ammonium transporters in Coffea
(Sociedade Brasileira de Genética, 2017) Santos, Tiago Benedito dos; Lima, Joni Esrom; Felicio, Mariane Silva; Soares, João Danillo Moura; Domingues, Douglas Silva
Nitrogen (N) is quantitatively the main nutrient required by coffee plants, with acquisition mainly by the roots and mostly exported to coffee beans. Nitrate (NO3 –) and ammonium (NH4 +) are the most important inorganic sources for N uptake. Several N transporters encoded by different gene families mediate the uptake of these compounds. They have an important role in source preference for N uptake in the root system. In this study, we performed a genome-wide analysis, including in silico expression and phylogenetic analyses of AMT1, AMT2, NRT1/PTR, and NRT2 transporters in the recently sequenced Coffea canephora genome. We analyzed the expression of six selected transporters in Coffea arabica roots submitted to N deficiency. N source preference was also analyzed in C. arabica using isotopes. C. canephora N transporters follow the patterns observed for most eudicots, where each member of theAMT andNRT families has a particular role in N mobilization, and where some of these are modulated by N deficiency. Despite the prevalence of putative nitrate transporters in the Coffea genome, ammonium was the preferential inorganic N source for N-starved C. arabica roots. This data provides an important basis for fundamental and applied studies to depict molecular mechanisms involved in N uptake in coffee trees.
Leaf-associated bacterial microbiota of coffee and its correlation with manganese and calcium levels on leaves
(Sociedade Brasileira de Genética, 2018) Sousa, Leandro Pio de; Silva, Marcio José da; Mondego, Jorge Maurício Costa
Coffee is one of the most valuable agricultural commodities and the plants’ leaves are the primary site of infection for most coffee diseases, such as the devastating coffee leaf rust. Therefore, the use of bacterial microbiota that inhabits coffee leaves to fight infections could be an alternative agricultural method to protect against coffee diseases. Here, we report the leaf-associated bacteria in three coffee genotypes over the course of a year, with the aim to determine the diversity of bacterial microbiota. The results indicate a prevalence of Enterobacteriales in Coffea canephora, Pseudomonadales in C. arabica ‘Obatã’, and an intriguing lack of bacterial dominance in C. arabica ‘Catuaí’. Using PERMANOVA analyses, we assessed the association between bacterial abundance in the coffee genotypes and environmental parameters such as temperature, precipitation, and mineral nutrients in the leaves. We detected a close relationship between the amount of Mn and the abundance of Pseudomonadales in ‘Obatã’ and the amount of Ca and the abundance of Enterobacteriales in C. canephora. We suggest that mineral nutrients can be key drivers that shape leaf microbial communities.