Prunus persica apoplastic proteome analysis reveals candidate proteins involved in the resistance and defense against Taphrina deformans.

Taphrina deformans is the fungus responsible for the peach leaf curl disease. To gain insight into the molecular mechanisms involved in plant resistance and response to the fungus, apoplastic differentially abundant proteins (DAPs) in a resistant (DR) and/or in a susceptible genotype (FL) were identified after 12 and 96 h post inoculation (hpi) and compared to those at 0 hpi. The Prunus persica apoplastic proteome was assessed by LC-MS/MS analysis. Altogether 332 proteins were identified, and their molecular and biological functions were classified. In both genotypes, major changes occurred at 96 hpi when the fungus had achieved the filamentous form. However, at 96 hpi, DR exhibited a greater number of increased proteins than FL. DAPs were enriched in biotic stress response, with most of the proteins belonging to the pathogenesis related (PR)-type. PRs exhibited the greatest fold changes of induction in DR. While PRs acting on pathogen cell wall (PR2, PR3 and PR4) were increased in both susceptible and resistant genotypes, others were exclusively induced in DR, such as some isoforms of PR5, defensin and PR17. Proteins exclusively induced in DR upon T.deformans inoculation such as four berberine bridge enzymes, two snakins and a GDS-lipase were identified. Moreover, upon inoculation cuticle was thickened to a greater extent in DR than in FL. This work reveals the active role of the apoplast against T. deformans and not only contributes to the elucidation of responses involved in resistance to leaf curl disease but also improves the knowledge on peach defenses against pathogens.

Phenology and fruit set of peach trees cultivated in Dois Vizinhos, Paraná State, Brazil / Fenologia e frutificação efetiva de pessegueiros cultivados em Dois Vizinhos, Paraná

Understanding the dormancy phases and dynamics of commercial cultivars in the regions where they are grown allows for the implementation of different cultural practices and aids in the understanding of the temperature requirements for collaboration with genetic improvement programs. The objective of this study was to determine the budding, flowering, and fruit set rates of peach trees cultivated in the municipality of Dois Vizinhos, Southwestern Paraná, Brazil. The present work was developed using the collection of peach trees in the fruit growing sector of the Universidade Tecnológica Federal do Paraná. Sixteen peach cultivars were evaluated: Rubimel, Leonense, Coral, Marli, Charme, Riograndense, Douradão, Chimarrita, Granada, BR1, Bonão, and Eldorado (all planted in 2009), as well as Regalo, Kampai, Fascínio, and Zilli (planted in 2014). The experimental design was completely randomized with four replications of one plant each, in a 16 × 2 bifactorial arrangement (cultivars × year/harvest). Phenology evaluations were performed weekly by counting open (anthesis) and sprouted flower buds, and the fruit set rate was measured just before fruit thinning. Flowering (%), budding (%), and fruit set (%) rates were calculated. It was found that the year/harvest 2016 promoted the highest rates of sprouting and fruit set. The Bonão cultivar showed greater flowering intensity and regularity in the crop years evaluated (2016 and 2017). Coral and Kampai present fruit sets with greater intensities and regularities when compared to the other cultivars evaluated under the climatic condition of the municipality of Dois Vizinhos. The Riograndense cultivar showed greater irregularity in flowering and fruit production.(AU)

Compreender as fases e a dinâmica da dormência de cultivares comerciais na região em que essa está inserida possibilita a execução de diferentes práticas culturais e auxilia na compreensão dos requisitos de frio e calor exigidos pela mesma, colaborando principalmente com os programas de melhoramento genético. O objetivo deste trabalho foi determinar as taxas de brotação, floração e frutificação efetiva de pessegueiros sob as condições ambientais de cultivo no município de Dois Vizinhos, região Sudoeste do Paraná. O presente trabalho foi desenvolvido na coleção de pessegueiros do setor de Fruticultura da Universidade Tecnológica Federal do Paraná. Foram avaliadas 16 cultivares de pessegueiros, sendo 'Rubimel', 'Leonense', 'Coral, 'Marli', 'Charme', 'Riograndense', 'Douradão', 'Chimarrita', 'Granada', 'BR-1', 'Bonão' e 'Eldorado' implantadas em 2009 e 'Regalo', 'Kampai', 'Fascínio' e 'Zilli', implantadas em 2014. O delineamento experimental foi inteiramente casualizado com quatro repetições de uma planta cada, no arranjo bifatorial 16 x 2 (cultivares x anos/safra). As avaliações de fenologia foram realizadas semanalmente através da contagem de gemas florais abertas (antese) e brotadas, e a taxa de frutificação efetiva foi realizada imediatamente antes do raleio de frutos. Foram calculadas as taxas de floração (%), de brotação (%) e de frutificação efetiva (%). Nas condições climáticas do município de Dois Vizinhos, no Estado do Paraná, verificou-se que o ano/safra 2016 promoveu as maiores taxas de brotação e frutificação efetiva. A cultivar 'Bonão' apresentou maior intensidade e regularidade de floração nos anos safra avaliados (2016 e 2017). As cultivares 'Coral' e 'Kampai' apresentam frutificação efetiva com maior intensidades e regularidade em relação aos demais cultivares avaliados para a condição de clima do local avaliado. A cultivar 'Riograndense' apresentou maior irregularidade na floração e na produção de frutos.(AU)

Identifying sources of metabolomic diversity and reconfiguration in peach fruit: taking notes for quality fruit improvement.

The metabolomic content determines many of the important features of a fruit, such as its taste, flavor, color, nutritional value, and abiotic or biotic resistance. Peach (Prunus persica (L.) Batsch) is one of the best genetically characterized species used as a model for Rosaceae, the drupes of which are a source of minerals, vitamins, fiber, and antioxidant compounds for healthy diets around the world. During the last few years, a great advance in the analysis of the metabolic diversity and reconfiguration in different peach varieties in response to developmental and environmental factors has occurred. These studies have shown that the great phenotypic diversity among different peach varieties is correlated with differential metabolomic content. Besides, the fruit metabolome of each peach variety is not static; on the contrary, it is drastically configured in response to both developmental and environmental signals, and moreover, it was found that these metabolic reconfigurations are also variety dependent. In the present review, the main sources of metabolic diversity and conditions that induce modifications in the peach fruit metabolome are summarized. It is postulated that comparison of the metabolic reconfigurations that take place among the fruits from different varieties may help us better understand peach fruit metabolism and their key drivers, which in turn may aid in the future design of high-quality peach fruits.

Fine-tuning the performance of ddRAD-seq in the peach genome.

The advance of Next Generation Sequencing (NGS) technologies allows high-throughput genotyping at a reasonable cost, although, in the case of peach, this technology has been scarcely developed. To date, only a standard Genotyping by Sequencing approach (GBS), based on a single restriction with ApeKI to reduce genome complexity, has been applied in peach. In this work, we assessed the performance of the double-digest RADseq approach (ddRADseq), by testing 6 double restrictions with the restriction profile generated with ApeKI. The enzyme pair PstI/MboI retained the highest number of loci in concordance with the in silico analysis. Under this condition, the analysis of a diverse germplasm collection (191 peach genotypes) yielded 200,759,000 paired-end (2 × 250 bp) reads that allowed the identification of 113,411 SNP, 13,661 InDel and 2133 SSR. We take advantage of a wide sample set to describe technical scope of the platform. The novel platform presented here represents a useful tool for genomic-based breeding for peach.

Shotgun proteomics of peach fruit reveals major metabolic pathways associated to ripening.

BACKGROUND: Fruit ripening in Prunus persica melting varieties involves several physiological changes that have a direct impact on the fruit organoleptic quality and storage potential. By studying the proteomic differences between the mesocarp of mature and ripe fruit, it would be possible to highlight critical molecular processes involved in the fruit ripening. RESULTS: To accomplish this goal, the proteome from mature and ripe fruit was assessed from the variety O'Henry through shotgun proteomics using 1D-gel (PAGE-SDS) as fractionation method followed by LC/MS-MS analysis. Data from the 131,435 spectra could be matched to 2740 proteins, using the peach genome reference v1. After data pre-treatment, 1663 proteins could be used for comparison with datasets assessed using transcriptomic approaches and for quantitative protein accumulation analysis. Close to 26% of the genes that code for the proteins assessed displayed higher expression at ripe fruit compared to other fruit developmental stages, based on published transcriptomic data. Differential accumulation analysis between mature and ripe fruit revealed that 15% of the proteins identified were modulated by the ripening process, with glycogen and isocitrate metabolism, and protein localization overrepresented in mature fruit, as well as cell wall modification in ripe fruit. Potential biomarkers for the ripening process, due to their differential accumulation and gene expression pattern, included a pectin methylesterase inhibitor, a gibbellerin 2-beta-dioxygenase, an omega-6 fatty acid desaturase, a homeobox-leucine zipper protein and an ACC oxidase. Transcription factors enriched in NAC and Myb protein domains would target preferentially the genes encoding proteins more abundant in mature and ripe fruit, respectively. CONCLUSIONS: Shotgun proteomics is an unbiased approach to get deeper into the proteome allowing to detect differences in protein abundance between samples. This technique provided a resolution so that individual gene products could be identified. Many proteins likely involved in cell wall and sugar metabolism, aroma and color, change their abundance during the transition from mature to ripe fruit.

Genome-wide SNP identification in Prunus rootstocks germplasm collections using Genotyping-by-Sequencing: phylogenetic analysis, distribution of SNPs and prediction of their effect on gene function.

Genotyping-by-Sequencing (GBS) was applied in a set of 53 diploid Prunus rootstocks and five scion cultivars from three subgenera (Amygdalus, Prunus and Cerasus) for genome-wide SNP identification and to assess genetic diversity of both Chilean and Spanish germplasm collections. A group of 45,382 high quality SNPs (MAF >0.05; missing data <5%) were selected for analysis of this group of 58 accessions. These SNPs were distributed in genic and intergenic regions in the eight pseudomolecules of the peach genome (Peach v2.0), with an average of 53% located in exonic regions. The genetic diversity detected among the studied accessions divided them in three groups, which are in agreement with their current taxonomic classification. SNPs were classified based on their putative effect on annotated genes and KOG analysis was carried out to provide a deeper understanding of the function of 119 genes affected by high-impact SNPs. Results demonstrate the high utility for Prunus rootstocks identification and studies of diversity in Prunus species. Also, given the high number of SNPs identified in exonic regions, this strategy represents an important tool for finding candidate genes underlying traits of interest and potential functional markers for use in marker-assisted selection.

Generation of fruit postharvest gene datasets and a novel motif analysis tool for functional studies: uncovering links between peach fruit heat treatment and cold storage responses.

MAIN CONCLUSION: A survey of developed fruit gene-specific datasets and the implementation of a novel cis-element analysis tool indicate specific transcription factors as novel regulatory actors under HT response and CI protection. Heat treatment (HT) prior to cold storage (CS) has been successfully applied to ameliorate fruit chilling injury (CI) disorders. Molecular studies have identified several HT-driven benefits and putative CI-protective molecules and mechanisms. However, bioinformatic tools and analyses able to integrate fruit-specific information are necessary to begin functional studies and breeding projects. In this work, a HT-responsive gene dataset (HTds) and four fruit expression datasets (FEds), containing gene-specific information from several species and postharvest conditions, were developed and characterized. FEds provided information about HT-responsive genes, not only validating their sensitivity to HT in different systems but also revealing most of them as CS-responsive. A special focus was given to peach heat treatment-sensitive transcriptional regulation by the development of a novel Perl motif analysis software (cisAnalyzer) and a curated plant cis-elements dataset (PASPds). cisAnalyzer is able to assess sequence motifs presence, localization, enrichment and discovery on biological sequences. Its implementation for the enrichment analysis of PASPds motifs on the promoters of HTds genes rendered particular cis-elements that indicate certain transcription factor (TF) families as responsible of fruit HT-sensitive transcription regulation. Phylogenetic and postharvest expression data of these TFs showed a functional diversity of TF families, with members able to fulfil roles under HT, CS and/or both treatments. All integrated datasets and cisAnalyzer tool were deposited in FruitGeneDB (https://www.cefobi-conicet.gov.ar/FruitGeneDB/search1.php), a new available database with a great potential for fruit gene functional studies, including the markers of HT and CS responses whose study will contribute to unravel HT-driven CI-protection and select tolerant cultivars.

Estimates of genetic parameters, genetic variability, and selection in the S1 generation of peach / Estimativas de parâmetros genéticos, variabilidade genética e seleção em geração S1 de pessegueiros

The growing of peach in mild winter regions is made viable through the use of genotypes that have low need for cold conditions, and this is one of the main aims of breeding for these regions. Thus, the aims of this study were to estimate genetic parameters, evaluate genetic variability, and select families adapted to mild winter regions in the S1 generation of peach through mixed model methodology (REML/BLUP). For that purpose, 22 populations, 84 families, and 2090 individuals were evaluated for the following traits: bud burst rate (BR), node density (ND), plant height (PH), and trunk diameter (TD). Genetic variability was found for all the traits. Individual heritability in the broad sense was of low and medium magnitudes. The PH trait had positive genotypic correlation of high magnitude with TD. The ND trait had moderate negative genotypic correlation with PH and TD. Clustering by the Tocher method resulted in the formation of six mutually exclusive groups. Considering selection intensity of 25%, simultaneous selection for BR, ND, and TD led to predicted gains of 11.3% for BR, 9.7% for ND, -14.2% for PH, and -14.3% for TD, showing the great potential of the germplasm evaluated.(AU)

O cultivo do pessegueiro em regiões de inverno ameno é viabilizado pela utilização de genótipos que apresentam baixa necessidade de frio, sendo este um dos principais objetivos do melhoramento para essas regiões. Assim, os objetivos deste estudo foram estimar parâmetros genéticos, avaliar a variabilidade genética e selecionar famílias adaptadas a regiões de inverno ameno em geração S1 de pessegueiros via metodologia de modelos mistos (REML/BLUP). Para isso, 22 populações, 84 famílias e 2090 indivíduos foram avaliados quanto as características: taxa de brotação (TB), densidade de nós (DN), altura da planta (AP) e diâmetro do tronco (DT). Verificou-se variabilidade genética para todas as características. As herdabilidades individuais no sentido amplo foram de baixa e média magnitudes. A característica AP apresentou correlação genética positiva de magnitude elevada com DT. A característica DN apresentou correlação genética negativa moderada com AP e DT. O agrupamento pelo método de Tocher resultou na formação de seis grupos mutuamente excludentes. Considerando intensidade de seleção de 25%, a seleção simultânea para TB, DN e DT propiciou ganhos preditos de 11.3% para TB, 9.7% para DN, -14.2% para AP e -14.3% para DT, evidenciando o grande potencial do germoplasma avaliado.(AU)

Qualidade e suscetibilidade de pêssegos e nectarinas aos danos causados pelo frio / Quality and susceptibility of peaches and nectarines to cold damage

Pêssegos e nectarinas são susceptíveis ao armazenamento refrigerado perdendo qualidade, assim a utilização de caracteres agronômicos correlacionados é importante em programas de melhoramento genético que visam frutos de qualidade e maior oferta da fruta no mercado. O objetivo deste estudo foi avaliar correlações entre os parâmetros de qualidade e de suscetibilidade aos danos por frio de pêssegos e nectarinas.O estudo foi conduzido na EEAD-CSIC, Zaragoza-Espanha, durante o ciclo produtivo 2013/2014. Um total de 40 acessos de pessegueiro e nectarineira foram avaliados quanto às características de qualidade [firmeza de polpa (FP), teor de sólidos solúveis totais (SST), acidez titulável (AT), pH, índice de maturação (IM), coloração do mesocarpo (CM)] e susceptibilidade aos danos por frio após armazenamento em duas temperaturas (0 e 5°C) e em dois períodos (14 e 28 dias). Dois dias após retirados das baixas temperaturas, foram observados diferentes sintomas de frio, como presença de polpa farinhenta (PF), granulosa (PG), coriácea (PC), alterações de coloração do mesocarpo, escurecimento (EI), avermelhamento interno (AI) e ausência de sabor (AS). Os dados foram submetidos ao estudo do coeficiente de correlação de Pearson entre os parâmetros de qualidade inicial e os frutos armazenados após duas e quatro semanas em 0 e 5°C. As variáveis IM, SST, FP e AT apresentaram correlações significativas entre si e, com PG e EI, indicando que frutos com maior amadurecimento tendem a desenvolver tais sintomas de danos. Os distúrbios EI, PF, PG e AS foram correlacionados significativamente, corroborando que esses sintomas são os principais distúrbios causados pelo armazenamento refrigerado. As variáveis de CM correlacionaram-se significativamente com EI e AIe,outros sintomas correlatos a estes, confirmando as alterações de cor. Palavras chave: Prunus pérsica L.; melhoramento genético; armazenamento refrigerado.

Peaches and nectarines are susceptible cold storage already it lost quality. Then, the use of related agronomic traits is important in breeding programs aimed to obtain fruit quality and higher fruit supply in the market. The aim of this work was to evaluate the correlations between quality and susceptibility to chilling injury of peaches and nectarines. The study was carried out in EEAD-CSIC, Zaragoza-Spain, during 2013/2014 cycle. A total of 40 peaches and nectarines genotypes from germplasm collection were evaluated. The quality characteristics as flesh firmness, total soluble solids, titratable acidity, pH, rippining index and flesh color parameters and the susceptibility chilling injury after storage in two temperatures (0 e 5 °C) and two times period (14 e 28 days) were evaluated. After two days that the fruits were removal these low temperatures, the presence of symptoms, such as wooliness through mealiness, flesh grainy, leatheriness and flesh color changes, through browning, bleeding and off flavor were evaluated. The data were submitted to study the Pearson’s correlation coefficient between the initial fruit quality variables and the fruits stored after two and four weeks at 0 and 5 °C. The quality parameters as rippining index, soluble solids, firmness and titratable acidity presented correlation among them. These, also it had correlation with woolines and bowning, what it indicated that fruits with more ripening can have the symptoms more easily. The browning, mealiness, flesh grainy and off flavor variables were correlationed with the time period and temperatures, what it confirms that these symptoms are the main disorders caused by cold storage. The flesh color variables presented significant correlations with browning and bleeding, and other symptoms related to, confirming color changes.

Qualidade e suscetibilidade de pêssegos e nectarinas aos danos causados pelo frio / Quality and susceptibility of peaches and nectarines to cold damage

Pêssegos e nectarinas são susceptíveis ao armazenamento refrigerado perdendo qualidade, assim a utilização de caracteres agronômicos correlacionados é importante em programas de melhoramento genético que visam frutos de qualidade e maior oferta da fruta no mercado. O objetivo deste estudo foi avaliar correlações entre os parâmetros de qualidade e de suscetibilidade aos danos por frio de pêssegos e nectarinas.O estudo foi conduzido na EEAD-CSIC, Zaragoza-Espanha, durante o ciclo produtivo 2013/2014. Um total de 40 acessos de pessegueiro e nectarineira foram avaliados quanto às características de qualidade [firmeza de polpa (FP), teor de sólidos solúveis totais (SST), acidez titulável (AT), pH, índice de maturação (IM), coloração do mesocarpo (CM)] e susceptibilidade aos danos por frio após armazenamento em duas temperaturas (0 e 5°C) e em dois períodos (14 e 28 dias). Dois dias após retirados das baixas temperaturas, foram observados diferentes sintomas de frio, como presença de polpa farinhenta (PF), granulosa (PG), coriácea (PC), alterações de coloração do mesocarpo, escurecimento (EI), avermelhamento interno (AI) e ausência de sabor (AS). Os dados foram submetidos ao estudo do coeficiente de correlação de Pearson entre os parâmetros de qualidade inicial e os frutos armazenados após duas e quatro semanas em 0 e 5°C. As variáveis IM, SST, FP e AT apresentaram correlações significativas entre si e, com PG e EI, indicando que frutos com maior amadurecimento tendem a desenvolver tais sintomas de danos. Os distúrbios EI, PF, PG e AS foram correlacionados significativamente, corroborando que esses sintomas são os principais distúrbios causados pelo armazenamento refrigerado. As variáveis de CM correlacionaram-se significativamente com EI e AIe,outros sintomas correlatos a estes, confirmando as alterações de cor. Palavras chave: Prunus pérsica L.; melhoramento genético; armazenamento refrigerado.(AU)

Peaches and nectarines are susceptible cold storage already it lost quality. Then, the use of related agronomic traits is important in breeding programs aimed to obtain fruit quality and higher fruit supply in the market. The aim of this work was to evaluate the correlations between quality and susceptibility to chilling injury of peaches and nectarines. The study was carried out in EEAD-CSIC, Zaragoza-Spain, during 2013/2014 cycle. A total of 40 peaches and nectarines genotypes from germplasm collection were evaluated. The quality characteristics as flesh firmness, total soluble solids, titratable acidity, pH, rippining index and flesh color parameters and the susceptibility chilling injury after storage in two temperatures (0 e 5 °C) and two times period (14 e 28 days) were evaluated. After two days that the fruits were removal these low temperatures, the presence of symptoms, such as wooliness through mealiness, flesh grainy, leatheriness and flesh color changes, through browning, bleeding and off flavor were evaluated. The data were submitted to study the Pearsons correlation coefficient between the initial fruit quality variables and the fruits stored after two and four weeks at 0 and 5 °C. The quality parameters as rippining index, soluble solids, firmness and titratable acidity presented correlation among them. These, also it had correlation with woolines and bowning, what it indicated that fruits with more ripening can have the symptoms more easily. The browning, mealiness, flesh grainy and off flavor variables were correlationed with the time period and temperatures, what it confirms that these symptoms are the main disorders caused by cold storage. The flesh color variables presented significant correlations with browning and bleeding, and other symptoms related to, confirming color changes.(AU)

Comparative Transcriptome Profiling in a Segregating Peach Population with Contrasting Juiciness Phenotypes.

Cold storage of fruit is one of the methods most commonly employed to extend the postharvest lifespan of peaches ( Prunus persica (L.) Batsch). However, fruit quality in this species is affected negatively by mealiness, a physiological disorder triggered by chilling injury after long periods of exposure to low temperatures during storage and manifested mainly as a lack of juiciness, which ultimately modifies the organoleptic properties of peach fruit. The aim of this study was to identify molecular components and metabolic processes underlying mealiness in susceptible and nonsusceptible segregants. Transcriptome and qRT-PCR profiling were applied to individuals with contrasting juiciness phenotypes in a segregating F2 population. Our results suggest that mealiness is a multiscale phenomenon, because juicy and mealy fruit display distinctive reprogramming processes affecting translational machinery and lipid, sugar, and oxidative metabolism. The candidate genes identified may be useful tools for further crop improvement.

A Prunus persica genome-wide RNA-seq approach uncovers major differences in the transcriptome among chilling injury sensitive and non-sensitive varieties.

Chilling injury represents a major constrain for crops productivity. Prunus persica, one of the most relevant rosacea crops, have early season varieties that are resistant to chilling injury, in contrast to late season varieties, which display chilling symptoms such as mealiness (dry, sandy fruit mesocarp) after prolonged storage at chilling temperatures. To uncover the molecular processes related to the ability of early varieties to withstand mealiness, postharvest and genome-wide RNA-seq assessments were performed in two early and two late varieties. Differences in juice content and ethylene biosynthesis were detected among early and late season fruits that became mealy after exposed to prolonged chilling. Principal component and data distribution analysis revealed that cold-stored late variety fruit displayed an exacerbated and unique transcriptome profile when compared to any other postharvest condition. A differential expression analysis performed using an empirical Bayes mixture modeling approach followed by co-expression and functional enrichment analysis uncover processes related to ethylene, lipids, cell wall, carotenoids and DNA metabolism, light response, and plastid homeostasis associated to the susceptibility or resistance of P. persica varieties to chilling stress. Several of the genes related to these processes are in quantitative trait loci (QTL) associated to mealiness in P. persica. Together, these analyses exemplify how P. persica can be used as a model for studying chilling stress in plants.

Construction of a highly saturated linkage map in Japanese plum (Prunus salicina L.) using GBS for SNP marker calling.

This study reports the construction of high density linkage maps of Japanese plum (Prunus salicina Lindl.) using single nucleotide polymorphism markers (SNPs), obtained with a GBS strategy. The mapping population (An x Au) was obtained by crossing cv. "Angeleno" (An) as maternal line and cv. "Aurora" (Au) as the pollen donor. A total of 49,826 SNPs were identified using the peach genome V2.1 as a reference. Then a stringent filtering was carried out, which revealed 1,441 high quality SNPs in 137 An x Au offspring, which were mapped in eight linkage groups. Finally, the consensus map was built using 732 SNPs which spanned 617 cM with an average of 0.96 cM between adjacent markers. The majority of the SNPs were distributed in the intragenic region in all the linkage groups. Considering all linkage groups together, 85.6% of the SNPs were located in intragenic regions and only 14.4% were located in intergenic regions. The genetic linkage analysis was able to co-localize two to three SNPs over 37 putative orthologous genes in eight linkage groups in the Japanese plum map. These results indicate a high level of synteny and collinearity between Japanese plum and peach genomes.

Comparative proteomic and metabolomic studies between Prunus persica genotypes resistant and susceptible to Taphrina deformans suggest a molecular basis of resistance.

The worldwide-distributed leaf peach curl disease is caused by the biotroph Taphrina deformans. To characterize the plant-fungus interaction, resistant and susceptible Prunus persica genotypes grown in the orchard were studied. Asymptomatic leaves were tested for fungal presence. In all resistant leaves analyzed the fungus was not detected. Conversely, leaves from the susceptible genotype were categorized according to the presence or absence of the pathogen. Comparative metabolomic analysis disclosed the metabolite composition associated with resistant and susceptible interactions, and of compounds involved in fungal growth inhibition such as chlorogenic acid, whose in vitro antifungal activity was verified in this work. Differential proteome studies revealed that chloroplasts are important site of plant defense responses against T. deformans. Members of the Bet-v1-like family protein differentially responded to the pathogen. Extracellular pathogenesis-related proteins, evaluated by qRT-PCR, and an enone oxidoreductase are constitutively present in leaves of resistant trees and could be related to fungal resistance. This study is a global view of the changes in the metabolome, proteome and transcripts related to plant defense in naturally infected leaves of susceptible plants during the asymptomatic stage. Additionally, it provides clues to the successful molecular mechanisms operating in resistant plants, which neither develop the disease nor harbor the pathogen.

Identification of reference genes for RT-qPCR analysis in peach genotypes with contrasting chilling requirements.

Selecting and validating reference genes are the first steps in studying gene expression by reverse transcriptase-quantitative polymerase chain reaction (RT-qPCR). The present study aimed to evaluate the stability of five reference genes for the purpose of normalization when studying gene expression in various cultivars of Prunus persica with different chilling requirements. Flower bud tissues of nine peach genotypes from Embrapa's peach breeding program with different chilling requirements were used, and five candidate reference genes based on the RT-qPCR that were useful for studying the relative quantitative gene expression and stability were evaluated using geNorm, NormFinder, and bestKeeper software packages. The results indicated that among the genes tested, the most stable genes to be used as reference genes are Act and UBQ10. This study is the first survey of the stability of reference genes in peaches under chilling stress and provides guidelines for more accurate RT-qPCR results.

Unravelling early events in the Taphrina deformans-Prunus persica interaction: an insight into the differential responses in resistant and susceptible genotypes.

Leaf peach curl is a devastating disease affecting leaves, flowers and fruits, caused by the dimorphic fungus Taphrina deformans. To gain insight into the mechanisms of fungus pathogenesis and plant responses, leaves of a resistant and two susceptible Prunus persica genotypes were inoculated with blastospores (yeast), and the infection was monitored during 120 h post inoculation (h.p.i.). Fungal dimorphism to the filamentous form and induction of reactive oxygen species (ROS), callose synthesis, cell death and defence compound production were observed independently of the genotype. Fungal load significantly decreased after 120 h.p.i. in the resistant genotype, while the pathogen tended to grow in the susceptible genotypes. Metabolic profiling revealed a biphasic re-programming of plant tissue in susceptible genotypes, with an initial stage co-incident with the yeast form of the fungus and a second when the hypha is developed. Transcriptional analysis of PRs and plant hormone-related genes indicated that pathogenesis-related (PR) proteins are involved in P. persica defence responses against T. deformans and that salicylic acid is induced in the resistant genotype. Conducted experiments allowed the elucidation of common and differential responses in susceptible versus resistant genotypes and thus allow us to construct a picture of early events during T. deformans infection.

Cloning and bioinformatic analysis of transcription factor MYB10 from the red-leaf peach.

In higher plants, the transcription factor MYB10 is an important regulator of anthocyanin biosynthesis. In order to study its role in the development of red coloration in peach leaves, the full-length MYB10 complementary DNA sequence of the red-leaf peach cultivar 'Tsukuba No. 5' (Prunus persica f. atropurpurea) was successfully cloned using reverse transcription-polymerase chain reaction. The sequence was assigned the GenBank accession No. KP315904. Bioinformatic analysis identified the complete MYB10 open reading frame, consisting of 678 bp encoding 225 amino acids. The predicted protein has a molecular weight of 26.56 kDa and a theoretical isoelectric point of 8.97. The secondary structure was found to comprise approximately 34.22% alpha helix, 15.11% extended strand, 10.67% beta turn, and 40% random coil. Subcellular analysis indicated that MYB10 may function in the cytoplasm. Assessment of the amino acid sequence suggested the presence of one serine and two threonine phosphorylation sites. Quantitative real-time polymerase chain reaction revealed that MYB10 expression positively correlated with anthocyanin content in red-leaf peach, indicating that this transcription factor plays a role in the biosynthesis of this pigment in peach trees.

Cell wall-related genes studies on peach cultivars with differential susceptibility to woolliness: looking for candidates as indicators of chilling tolerance.

KEY MESSAGE: The results obtained indicate that a ß-xylosidase gene may act as good indicator of chilling tolerance and provide new insights into the complex issue of peach fruit woolliness. The storage of peaches at low temperatures for prolonged periods can induce a form of chilling injury (CI) called woolliness, characterized by a lack of juiciness and a mealy texture. As this disorder has been associated with abnormal cell wall dismantling, the levels of 12 transcripts encoding proteins involved in cell wall metabolism were analysed in cultivars with contrasting susceptibility to this disorder selected from five melting flesh peach cultivars. The resistant ('Springlady') and susceptible ('Flordaking') cultivars displayed differences in the level of expression of some of the selected genes during fruit softening and in woolly versus non-woolly fruits. From these genes, the level of expression of PpXyl, which encodes for a putative ß-xylosidase, was the one that presented the highest correlation (negative) with the susceptibility to woolliness. PpXyl expression was also analysed in a cultivar ('Rojo 2') with intermediate susceptibility to woolliness, reinforcing the conclusion about the correlation of PpXyl expression to the presence of woolliness symptom. Moreover, the level of expression of PpXyl correlated to protein level detected by Western blot. Analyses of the promoter region of the PpXyl gene (1637 bp) isolated from the three cultivars showed no differences suggesting that cis-elements from other regions of the genome and/or trans elements could be responsible of the differential PpXyl expression patterns. Overall, the results obtained indicate that PpXyl may act as a good indicator of woolliness tolerance and that the regulation of expression of this gene in different cultivars does not depend on sequences upstream the coding sequence.

Transcriptome analysis of peach (Prunus persica L. Batsch) during the late stage of fruit ripening.

Fruit ripening is a complex developmental process, the details of which remain largely unknown in fleshy fruits. In this paper, the fruit flesh of two peach varieties, "Zhongyou9" (a nectarine; Prunus persica L. Batsch) and its mutant "Hongyu", was analyzed by RNA-seq technology during two stages of ripening at 20-day intervals. One hundred and eighty significant upregulated and two hundred and thirty-five downregulated genes were identified in the experiment. Many of these genes were related to plant hormones, chlorophyll breakdown, accumulation of aroma and flavor volatiles, and stress. To the best of our knowledge, this is the first transcriptome analysis of peach ripening, and our data will be useful for further studies of the molecular basis of fruit ripening.

Analysis of biochemical compounds and differentially expressed genes of the anthocyanin biosynthetic pathway in variegated peach flowers.

Variegated plants are highly valuable in the floricultural market, yet the genetic mechanism underlying this attractive phenomenon has not been completely elucidated. In this study, we identified and measured different compounds in pink and white flower petals of peach (Prunus persica) by high-performance liquid chromatography and liquid chromatography/mass spectrometry analyses. No cyanidin-based or pelargonidin-based compounds were detected in white petals, but high levels of these compounds were found in pink petals. Additionally, we sequenced and analyzed the expression of six key structural genes in the anthocyanin biosynthesis pathway (CHI, CHS, DFR, F3'H, ANS, and UFGT) in both white and pink petals. Quantitative real-time polymerase chain reaction revealed all six genes to be expressed at greatly reduced levels in white flower petals, relative to pink. No allelic variations were found in the transcribed sequences. However, alignment of transcribed and genomic sequences of the ANS gene detected alternative splicing, resulting in transcripts of 1.071 and 942 bp. Only the longer transcript was observed in white flower petals. Since ANS is the key intermediate enzyme catalyzing the colorless leucopelargonidin and leucocyanidin to substrates required for completion of anthocyanin biosynthesis, the ANS gene is implicated in flower color variegation and should be explored in future studies. This article, together with a previous transcriptome study, elucidates the mechanism underlying peach flower color variegation in terms of the key structural genes involved in anthocyanin biosynthesis.