Research Article

Table of Contents | Genet. Mol. Res. 2018 (4)

Displaying 1 - 15 of 20
Research Article

Some species of Anacardium (Anacardiaceae) produce fruits and pseudofruits that are smaller than those of the common cashew (Anacardium occidentale) and, for this reason, are known collectively in Brazil as “cajuí”. Despite their economic value in the food market and their important environmental and ecological functions, cajuí trees remain underexploited. We employed nine inter-simple sequence repeat (ISSR) markers to characterize two presupposed populations of cajuí comprising 25 accessions maintained in the germplasm bank of Embrapa Meio-Norte (Teresina, PI, Brazil). Population structure and relationships between accessions were determined in order to generate knowledge that could contribute to genetic improvement programs and better management of this germplasm bank. A high degree of polymorphism (91.3%) was observed among the accessions. Analysis of molecular variance and Bayesian analysis demonstrated that the two presupposed populations were not genetically differentiated but constituted a single population containing highly diversified individuals including migrants, migrant descendants and possible hybrids. Nonetheless, genetic variability within the accessions could be organized into two distinct, but linked, groups that had undergone extensive exchange of genetic material, as verified by the high gene flow index ). The substantial genetic variability observed could be attributed to individual differences between accessions rather than to differential spatial distribution. This report enhances our knowledge of the genus Anacardium and should facilitate the future improvement of cajuí culture and fruit quality. In addition, our study highlights the importance of further taxonomic studies on the species of Anacardium that comprise cajuí.

Genet. Mol. Res. 17(4): GMR18212
DOI: 10.4238/gmr18212
Research Article

The oleaginous species Jatropha curcas (Euphorbiaceae) exhibits significant potential as a source of biodiesel, but the scarcity of information regarding genetic variability within the species limits its possible exploitation. We examined the genetic diversity and relationships among 97 accessions of J. curcas originating from the Brazilian states of Piauí, Maranhão, Ceará, Bahia, and Minas Gerais. One-hundred ISSR markers were tested and 11 selected for genotyping. Among 307 loci generated by these markers, 96 % were polymorphic. Genetic similarities between accessions were estimated from Jaccard coefficients and the corresponding similarity matrix, and genetic relationships were determined from the dendrogram constructed using the unweighted pair group method with arithmetic average (UPGMA) technique. Nei’s genetic diversity indices varied between 0.0256and 0.1703, while Shannon’s information indices ranged from 0.0374 to 0.1926. The number of alleles varied from 1.0619 to 1.3257 and the effective number of alleles ranged between 1.0438 and 1.1496. The inter-population genetic coefficient differentiation was 0.6306, while analysis of molecular variance revealed that genetic divergence among populations was highly significant (p< 0.001), expressed by a fixation index (ΦST) of 0.4452. Pair-wise analysis of ΦST confirmed an inter-population diversity of 44.52% and an intra-population variation of 55.48%. UPGMA analysis allowed the separation of the accessions into four genotypic groups. We conclude that there is significant genetic diversity among the populations of J. curcas and that this variability is mainly due to intra-population genetic diversity.

 

Genet. Mol. Res. 17(4): GMR18208
DOI: 10.4238/gmr18208
Research Article

The common bean (Phaseolus vulgaris) is a widespread crop in Brazil of dietary and economic importance, and it is cultivated primarily through family farming. Knowledge of genetic variability in landraces and improved bean cultivars is essential to explore the existing diversity, identify superior genotypes adapted to the climatic conditions of specific regions, and support genetic improvement strategies. Estimates of genetic diversity can be obtained using DNA molecular markers, and ISSR markers are widely used. We evaluated the genetic diversity of 57 common bean genotypes, including accessions provided by the Brazilian Agricultural Research Corporation (EMBRAPA - Wheat), local genotypes of the Fortaleza community (Muqui-Espírito Santo) and commercial cultivars, using ISSR markers. A total of 11 primers were used, generating 51 fragments, of which 76%were polymorphic. The polymorphic information content ranged from 0.19 to 0.48, with a mean of 0.36. There was an unequal distribution between genetic distances, ranging from 0.00 to 1.0, and a mean of 0.44, evidencing wide genetic variability. The Pérola cultivar stood out as it showed the highest mean dissimilarity (0.76). Cluster analysis revealed the formation of 11 groups, with a tendency to cluster genotypes by the region of origin and growth habit. There was wide genetic diversity among the genotypes of the Fortaleza community and a narrower diversity for the EMBRAPA and commercial cultivars. ISSR markers were efficient in quantifying the genetic diversity of the genotypes; the most divergent markers will help select candidates for conservation in germplasm banks.

 

Genet. Mol. Res. 17(4): GMR18106
DOI: 10.4238/gmr18106
Research Article

Soil water limitations can cause high losses in agricultural yields. In order to investigate how popcorn varieties are affected by reduced water availability, we evaluated grain yield (GY) and popping expansion (PE) of 20 popcorn lines under water stress (WS) and well-watered conditions (WW), to propose discrimination with regard to the level of water-stress tolerance (T) and agronomic water-use efficiency (WUE), as well as to identify crosses for the breeding of superior hybrid combinations and for inheritance studies. A randomized complete block design with three replications was used. Irrigation was applied at pre-anthesis. The germplasm was discriminated based on the Stress tolerance index (STI), Stress susceptibility index (SSI), Stability index (SIN), Drought resistance index (DRI), and Agronomic water-use efficiency (WUE). Genetic diversity was measured by 15 EST-SSR markers. The reduction in GY under water stress was 55.29% and PE was reduced 29.19%. For the identification of genotypes with higher phenotypic means, STI and WUE were similarly efficient, whereas SSI and SIN identified genotypes with a lower proportional performance loss in the WS compared to the WW environment. For both GY and PE, there was a lack of relationship between WUE (more productive) and T (more stable). To explore the allelic complementarity for WUE, for T and for both, respectively, the combinations L59 x P7, L55 x P1 and L71 x P6 had the best performances. The lines L61, L63 and L65 phenotypically contrasted to those with high WUE and T and could be used for inheritance studies. Genotypes with higher WUE are considered the most appropriate option for breeding programs under WS.

 

Genet. Mol. Res. 17(4): GMR18184
DOI: 10.4238/gmr18184
Research Article

Anopheles (Nyssorhynchus) darlingi is the primary vector of human malaria in South America. Immune responses in mosquito vectors of malaria are mainly regulated by genes of the Toll and IMD pathways through the transcription factors NF-kappa-β, Rel1 and Rel2, which are controlled by the negative regulatory genes Cactus and Caspar.  We measured the expression levels of Rel1, Rel2, Caspar and Cactus genes, which are related to the immune system, in adult females of A. darlingi after blood feeding compared to adult females without blood feeding (controls) due to their possible effects on the ability of becoming infected with species of Plasmodium and spreading malaria. Quantitative expression was determined by real-time PCR, using the reference genes GAPDH and β-actin. The expression levels of Rel1, Rel2, Caspar and Cactus varied significantly at 4, 8, 14 and 24 h in mosquitoes that had fed on blood compared to control insects (0 h), with significantly greater expression at 24 h after blood feeding. Relative expression levels among these genes varied at the different post blood feeding times. This information adds to our understanding of the insect immune response system and related questions involved in understanding the biology and control of this mosquito.

Genet. Mol. Res. 17(4): GMR18076
DOI: 10.4238/gmr18076
Research Article

The plants Brosimum gaudichaudii (inharé) and Caesalpinia ferrea (jucá) are widely distributed throughout Brazil and are considered medicinal. Inharé has been popularly used as a blood purifier, for the treatment of skin and vitiligo, while jucáis considered analgesic, anti-inflammatory and anticancer. These therapeutic properties have been attributed to phytochemicals such as coumarins, flavonoids and tannins. However, the mechanisms of action of most of these phytochemicals have not yet been fully elucidated and they may compromise human health. Consequently, the evaluation of the genotoxic effect of these plant extracts is fundamental for the determination of safe doses for human consumption. We evaluated the genotoxic effect of various concentrations of extracts of B. gaudichaudii and C. ferrea using comet assay of erythrocytes from Astyanax sp. exposed in vivo. In the comet assay indicated the tail migration of DNA increased significantly in the group of cells exposed to C. ferrea for various treatments and the olive tail movement exhibited a significantly higher extent of DNA damage, indicating the potential genotoxicity of the extract. On the other hand, it is premature to claima lack of genotoxic effect for B. gaudichauddi extracts since our experimental design was not able to rule out apotential effect of DNA damage as the concentration of 20mg/L seemed to increase the likelihood of genotoxicity. Thus, larger doses of B. gaudichauddi extracts should be tested in future studies of the kind. Our investigation provided valuable data for two species of plants widely used in folk medicine in different regions of Brazil. We recommend caution when using thses species for their medicinal properties.

Genet. Mol. Res. 17(4): GMR18100
DOI: 10.4238/gmr18100
Research Article

Non-steroidal anti-inflammatory drugs (NSAIDs) are widely-used medications for the treatment of many inflammatory diseases. . They inhibit cyclooxygenase (COX) enzymes as a primary mechanism of action. However, many of these drugs were withdrawn from the market because of cardiovascular side effects. To date, only a limited number of genes and specific gene variants that account for their cardiac toxicity have been investigated. To identify possible cardiotoxic effects, we performed a microarray analysis of NSAIDs-treated primary cardiomyocytes. Primary cultures of neonatal rat cardiomyocytes were established and a DNA microarray was run on extracted RNA from celecoxib and diclofenac-treated cardiomyocytes. The changes in gene expression for all potentially predisposing variants were investigated. In cells treated with celecoxib, the expression of 1718 and 560 genes decreased and increased respectively by 2-fold or more. When cells were treated with diclofenac, the expression of 424 and 32 genes decreased and increased respectively by 2-fold or more. NSAIDs affected the expression of genes involved in calcium and potassium signaling. Pathway analysis of gene expression of NSAIDs-treated cardiomyocytes showed changes in gene expression involving major pathways, including apoptosis, signal transduction and transcription. These findings provide a clue to explain, at least in part, how NSAIDs provoke side effects in the heart. These data could be used to elect target genes for studying cardiotoxic effects of NSAIDs and for developing new drugs.

 

Genet. Mol. Res. 17(4): GMR18068
DOI: 10.4238/gmr18068
Research Article

Plant transformation is a widely used procedure for obtaining transgenic plants and to develop plant models to understand gene function. Plant models such as Nicotiana tabacum are widely used for understanding gene responses to external influences. An important tool in such studies is genetic transformation through infection with Agrobacterium tumefaciens. However, this transformation is often inefficient. Consequently, development and optimization of techniques to promote high rates of seedling regeneration of transgenic tobacco is imperative. The methods tested for infection of tobacco explants consisted of injecting 10 μl of the bacterial culture directly into anodal segment using an insulin syringe (1 mL); bacterial co-cultivation with nodal segments and micro-sectioned leaf disks. Infection through punctures made with a syringe in nodal segments of tobacco and no co-cultivation period was the most efficient in the regeneration process and in obtaining genetically transformed plants, with 88 and 75% success rates, respectively. We obtained an increase of 50% in the transformation rates when compared to previous studies using N. tabacum.

 

Genet. Mol. Res. 17(4): GMR18072
DOI: 10.4238/gmr18072
Research Article

Gold nanoparticles (GNPs) with controlled geometric properties are new options for the management of inflammatory responses and therapy for various diseases, including diabetes mellitus, rheumatoid arthritis, and connective tissue diseases. We examined anti-inflammatory impacts of sugar coated GNPs with different sizes on carrageenan-induced rat paw inflammation. The synthesized GNPs were compared with indomethacin, methotrexate and the commercial gold preparation myochrysine. While methotrexate and myochrysine did not have any significant effects on rat paws, GNPs significantly decreased the inflammatory responses, comparable with indomethacin, a standard anti-inflammatory drug in the rat paw model. There was no harmful effect on metabolic activity of cells treated with GNPs of different sizes. We conclude that sugar coated GNPs are a promising candidate for new options in the management of inflammatory-related diseases.

 

Genet. Mol. Res. 17(4): GMR18037
DOI: 10.4238/gmr18037
Research Article

Selection of appropriate genitors in breeding programs increases gains due to the variability found in the divergent groups; this allows quantification of the existing variability, saving time and resources. There are many methods for quantification and evaluation of diversity in population studies, among which we highlight methods that are based on multivariate statistical analyses, such as linear discriminant analysis (LDA) and cluster analysis. Here we propose and evaluate the use of Support Vector machine (SVM) and Artificial Neural Network (ANN) in an attempt to solve the problem of genetic classification of hybrid populations with high degrees of similarity. The results obtained, in terms of the apparent error rate (APER), were compared with those obtained using ANN analysis and LDA. In general, the lowest APER values were associated with scenarios with low degrees of genetic similarity between populations. Specifically, the best results obtained through SVM (ranging from 14.44 to 67.41%) were observed when the exponential radial base kernel function was used. The APERs obtained by the ANN were even lower than those of the linear discriminant function.

 

Genet. Mol. Res. 17(4): GMR18122
DOI: 10.4238/gmr18122
Research Article

Sine oculis-related homeobox 1 (Six1) and 4 (Six4) transcription factors are expressed in developing and adult muscle. These Six homeoproteins are required to activate the fast-type muscle program in the mouse primary myotome. Whether Six1 and Six4 genes have a role in meat tenderness in Tan sheep has not yet been established. To study the characteristics and mechanisms of meat tenderness in Tan sheep, we sampled the musculus triceps muscle, the longest neck muscles, abdominal muscles, gluteus and biceps femoris muscle and measured Warner-Bratzler shear force and texture profile analysis using a texture analyzer (TMS-Pro), and mRNA expression of Six1 and Six4by real-time PCR, and we performed a correlation analysis. The Warner-Bratzler shear force was significantly different among the five different muscles, but there was no difference in this measure between the triceps and biceps femoris muscles. Hardness, gumminess and chewiness were significantly different among the different muscles. Six1 and Six4 mRNA expression was significantly higher in the longest neck muscles than in the other muscles. There was a significant negative correlation between Six1 mRNA expression and the texture parameters gumminess and chewiness, and a significant positive correlation between Six4 mRNA expression and gumminess and chewiness. Based on these results, we suggest that Six1 and Six4 not only regulate muscle fiber typing, but also meat tenderness in Tan sheep. They could thus be candidate genes for selecting for meat tenderness.

 

Genet. Mol. Res. 17(4): GMR18098
DOI: 10.4238/gmr18098
Research Article

Zungo (ZU), San Pedreño (SP) and Casco de Mula (CM) are Colombian indigenous pig breeds originated from European populations that are well adapted to tropical conditions. These breeds have a great potential to be used in pure and crossbred schemes due to their high reproductive performance and resistance to tropical diseases. We investigated the genetic diversity and genetic structure of these pig breeds using microsatellite molecular markers. Fifty-five ZU, SP and CM animals were genotyped for 10 microsatellites using capillary electrophoresis. The average number of alleles per marker ranged from 5.56±1.88 in ZU to 6.70±1.64 in SP. Observed heterozygosity ranged from 0.68±0.05 in ZU to 0.74±0.03 in SP. Polymorphic information content was high for most of the microsatellites in all breeds. Inbreeding coefficient (FIS) values were low for all microsatellites, with an average of 0.035±0.037. Higher average values closer to and above 0.1 were observed for the fixation index of the global population (FIT) (0.131±0.041) and the coefficient of relatedness between individuals from different populations (FST) (0.099±0.013). Principal components and structure analyses showed a high level of clustering per breed and very low admixture levels between them. The panel of markers used in this study proved to be useful to investigate genetic diversity and pedigree relationships in pig populations. The Colombian pig breeds that we evaluated had a high genetic variability and a well-differentiated genetic structure, which are key for decision making in conservation programs and for the implementation of future animal breeding plans in these populations.

 

Genet. Mol. Res. 17(4): GMR18161
DOI: 10.4238/gmr18161
Research Article

The adenosine deaminase G22A polymorphism (20q.11.33) affects the level of adenosine deaminase (ADA) expression, which plays an important role in the regulation of intracellular and extracellular concentrations of adenosine.  Recent studies reported greater ADA activity in diabetic patients and showed the role of ADA in the modulation of insulin activity and glucose homeostasis. We investigated whether the G22A polymorphism of the ADA gene is associated with type 2 diabetes mellitus (T2DM) in the Palestinian population and assessed the relationship between the G22A variant and fasting plasma glucose (FPG), glycated hemoglobin (HbA1c) and lipid profile among T2DM patients.  A total of 231 individuals with T2DM and a control sample of 101 non-diabetic participants were randomly selected from those who were attending United Nations Relief and Works Agency (UNRWA) clinics for treatment and/or follow up.  Genomic DNA was extracted from peripheral blood samples and PCR-RFLP was performed to identify the TaqI polymorphism G22A of the ADA gene. No significant differences were observed in the genotype and allele frequencies between T2DM patients and the control group. Yet, among diabetic patients, the GG genotype was significantly associated with higher FPG and HbA1c when compared to the GA+AA genotype but had no influence on blood pressure, BMI or other metabolic parameters. In conclusion, we confirm that the GG genotype of the ADA gene is associated with poor glycemic control in T2DM Palestinians and points to the association of the G22A variant with decreased activity of the ADA enzyme, which is of paramount importance in the pathophysiology of T2DM.

 

Genet. Mol. Res. 17(4): GMR18111
DOI: 10.4238/gmr18111
Research Article

Morphological or isozyme markers related to physiological maturation and deteriorative processes are important in the evaluation of seed quality. Two experiments were conducted to examine the possibility of using isozymes as indicators of quality in tobacco seed lots and fruit appearance as an indicator of physiological maturity in tobacco cultivars, based on the physiological and biochemical changes of the seeds. Cultivars CSC 444 and CSC 221 of tobacco fruits were harvested at various maturity stages and their physiological quality was assessed by germination, first count, germination speed index, time to reach 50% germination, cumulative average germination, and seedling emergence. We also assessed the activity of catalase (EC 1.11.6.1 – CAT), esterase (EC 3.1.1.1 – EST), isocitrate dehydrogenase (EC 1.1.1.41 – IDH), malate dehydrogenase (EC 1.1.1.37 – MDH), alcohol dehydrogenase (EC 1.1.1.1 – ADH), endo-β-mannanase (EC 3.2.1.78), and heat-resistant proteins during the process of maturation. Six lots of cultivar CSC 444 were used to differentiate the quality levels between the lots, and their characterization was determined by germination and vigor tests. In addition, we evaluated the enzymatic activity of CAT, EST, ADH, MDH, and heat-resistant proteins. During maturation of the fruits from the partially dark stage, we observed a progressive increase in germination and seed vigor. We concluded that appearance of the fruit is an indicator of fruit maturity and quality in tobacco seeds. The enzymatic profile of ADH matches the physiological potential of the seeds, based on germination and emergence tests. Thus the ADH enzyme indicates the optimum stage to harvest fruits. In the EST and CAT enzymatic pattern analysis, we observed higher activity of these enzymes in lots with lower physiological quality. So the CAT and EST enzymes are biochemical indicators that can assess the deterioration of tobacco seed lots.

Genet. Mol. Res. 17(4): GMR18058
DOI: 10.4238/gmr18058
Research Article

Phosphate (Pi) unavailability is a growth-limiting factor for plants. Under Pi-limited conditions, plants activate molecular mechanisms for better acquisition and utilization of this nutrient. In maize, changes in the expression pattern of several Pi starvation-induced genes, including the A1 coding for dihydroflavonol 4-reductase (DFR) involved in anthocyanin biosynthesis, were identified through microarray analysis. In order to elucidate the molecular determinants with a potential role in P use efficiency, we carried out a study on gene expression analysis of the A1 phosphate responsive gene by northern blot analysis of total RNA from maize genotypes contrasting for Pi efficiency. Two Pi-efficient (L-03 and L-161-1) and five inefficient (L-11, L-16, L-22, L-53, and L-5046) genotypes of maize were grown for 15 days in hydroponic culture in the presence (250 mM Pi) or absence (0 mM Pi) of phosphate. All genotypes showed an increase in anthocyanin accumulation in roots in the absence of Pi (0 mM Pi). The Pi-efficient genotype L-36 and the Pi-inefficient genotypes L-16, L22, and L-5046 showed the highest levels of anthocyanin accumulation. The A1 gene exhibits temporal and spatial expression patterns associated with Pi deficiency. Although there were differences in the expression profile of Pi starvation induced genes, no consistent expression patterns could be associated with either Pi-efficient or Pi-inefficient genotypes. It appears that Pi efficiency in tropical maize is a complex trait mediated by a coordinated action of genes that are either induced or suppressed in response to Pi-deficiency.

 

Genet. Mol. Res. 17(4): GMR18036
DOI: 10.4238/gmr18036

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