Polinización y cuajado en ciruelo japonés

En algunos cultivares de ciruelo japonés se produce una caída masiva de flores y se obtienen porcentajes de cuajado muy bajos por causas que no están bien establecidas. Esta situación repercute negativamente en la cosecha y provoca grandes oscilaciones en la producción de un año a otro. En este trabajo se estudian las causas de los cuajados erráticos registrados en 12 cultivares de ciruelo japonés en condiciones reales de cultivo. Para ello, se han analizado el proceso de polinización y cuajado de frutos en campo y la incompatibilidad polen‐pistilo. Los ensayos de polinización controlada realizados en plantaciones comerciales con historiales de cuajados erráticos mostraron una clara influencia de la polinización en el cuajado en los cultivares analizados, ya que se obtuvo un cuajado mayor que el registrado en polinización libre o bien no se obtuvo cuajado, lo que indica que es necesaria una elección adecuada de cultivares polinizadores para obtener cuajado. La observación de la germinación del polen al microscopio ha permitido descartar la ausencia de insectos polinizadores y la falta de viabilidad del polen como las causas responsables de la falta de cuajado en las plantaciones estudiadas, aunque se han identificado dos nuevos cultivares androestériles, Black Jewell y Sybarite. Para determinar si la incompatibilidad polen‐pistilo está relacionada con las situaciones de falta de cuajado, se ha evaluado la auto(in)compatibilidad en un total de 30 de cultivares, incluyendo los de mayor importancia a nivel nacional, mediante la observación del crecimiento de tubos polínicos en flores autopolinizadas en campo y laboratorio. La combinación de ensayos de polinización y cuajado en campo, la determinación de la autoincompatibilidad mediante la observación del crecimiento de los tubos polínicos al microscopio y la identificación de los alelos S de incompatibilidad mediante PCR ha permitido identificar las causas de los bajos cuajados de 12 cultivares en 9 plantaciones comerciales. Uno de estos cultivares, Friar, se ha identificado como autocompatible, y los otros 11 como autoincompatibles (Angeleno, Blackamber, Black Diamond, Black Gold, Black Star, Earliqueen, Fortune, Golden Globe, Golden Japan, Larry Ann y Songold), para los que se han podido seleccionar cultivares polinizadores compatibles y coincidentes en floración en las condiciones de cultivo de cada plantación, lo que ha permitido aportar soluciones agronómicas a cada uno de las situaciones analizadas de falta de cuajado asociada a problemas de polinización. Se ha evaluado la auto(in)compatibilidad en otros 17 cultivares en flores autopolinizadas en laboratorio. Siete de ellos se han identificado como autocompatibles (Casselman, Laetitia, Nubiana, Rubirosa, Santa Rosa, Simka y Zanzi Sun) y 10 como autoincompatibles (Champion, Eldorado, Flavour Queen, Frontier, Green Sun, Laroda, Queen Ann, Queen Rosa, Sweet August y TC Sun). Se ha determinado también el genotipo S de un total de 115 cultivares, 97 de ellos descritos por primera vez, que incluyen la mayoría de los cultivados en la actualidad. Para ello se han utilizado técnicas moleculares ya establecidas en ciruelo japonés y se ha desarrollado un nuevo método, basado en la detección con electroforesis capilar, que han permitido distribuir los cultivares en sus correspondientes grupos de incompatibilidad y agilizar el proceso de selección de polinizadores. Los análisis moleculares de los genes del locus S en los cultivares analizados han permitido identificar cinco nuevos alelos de incompatibilidad (So‐Ss) y 14 nuevos grupos de incompatibilidad. Finalmente, en algunos cultivares se han detectado situaciones de falta de cuajado no relacionadas con problemas de polinización, sino con problemas en el desarrollo de los óvulos. Por un lado, en las flores emasculadas de algunos cruzamientos y, por otro, en plantaciones comerciales de dos nuevos cultivares (Rubirosa y Sweet August). En ambos casos la falta de cuajado se ha asociado a la degeneración prematura de los dos óvulos de la flor.In some Japanese plum cultivars a massive flower drop occurs and very low fruit set percentages are obtained for reasons that are not well established. This situation negatively influences crop load, causing large year‐to‐year variations in production. In this work, the causes of low and erratic fruit set of 12 Japanese plum cultivars are studied in commercial orchards. For this purpose, the pollination and fruit set processes in the field as well as the self‐incompatibility and ovule development have been analyzed. Controlled pollination assays carried out in commercial orchards with previous erratic fruit set records showed a clear influence of pollination on fruit set, because fruit set in controlled pollinations was higher or null than fruit set in open pollination conditions, thus revealing that most cultivars need an appropriate pollinator to produce fruit set. The observation of pollen germination under the microscope has allowed to discard the absence of pollinating insects and reduced pollen viability as the causes of lack of fruit set in the orchards studied, although two male sterile cultivars, Black Jewell and Sybarite were newly identified. To determine if self‐incompatibility is related to lack of fruit set, the self‐ (in)compatibility of 30 cultivars, including most relevant in Spanish orchards, was evaluated by the observation of pollen tube growth in self‐pollinated flowers in the field and the laboratory. The combined pollination and fruit set assays in the field, self‐incompatibility determination under the microscope and self‐incompatibility S allele identification by PCR have allowed identifying the causes of low fruit set in 12 cultivars from 9 commercial orchards. One of these cultivars, Friar, has been identified as self‐compatible, and the other 11 as self‐incompatible (Angeleno, Blackamber, Black Diamond, Black Gold, Black Star, Earliqueen, Fortune, Golden Globe, Golden Japan, Larry Ann and Songold), for which it was possible to select at least one compatible pollinator coincident in blooming time in the growing conditions of each orchard. This, it has been possible to provide agronomic solutions to each of the analyzed situations of lack of fruit set associated with pollination failure. The self(in)‐ compatibility has been evaluated in other 17 cultivars in self‐pollinated flowers in the laboratory. Seven of them were self‐compatible (Casselman, Laetitia, Nubiana, Rubirosa, Santa Rosa, Simka and Zanzi Sun) and 10 self‐incompatible (Champion,Eldorado, Flavour Queen, Frontier, Green Sun, Laroda, Queen Ann, Queen Rosa, Sweet August and TC Sun). The S genotype of 115 cultivars, 97 of them described for the first time, which include most of currently grown cultivars, was also identified. For this purpose, molecular techniques well established in Japanese plum were used and a new methodology, based on capillary electrophoresis detection, was developed. These results allowed assigning the cultivars analyzed to their corresponding incompatibility groups and expediting the process of pollinator selection. Molecular analysis of S‐locus genes in the cultivars tested allowed the identification of five new S‐alleles (So‐Ss) and 14 new incompatibility groups. Despite of these results, some cultivars showed lack of fruit set that was not related to pollination failure, but with abnormalities in the developmental process of ovules. In both emasculated flowers of some crosses, and in two new cultivars (Rubirosa and Sweet August), the lack of fruit was associated with premature degeneration of the two flower ovulesUnpublishe

Necesidades de polinización de las principales variedades de ciruelo japonés cultivadas en España

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Cerezo y ciruelo

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Evaluación del carácter de autocompatibilidad en nuevas variedades de ciruelo japonés (híbridos de Prunus salicina Lindl.): póster

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Extremadura y Aragón investigan la polinización del ciruelo japonés

El centro de Investigación Agraria Finca ‘La Orden- Valdesequera’ y el Centro de Investigación y Tecnología Agroalimentaria de Aragón (CITA) desarrollan conjuntamente proyectos de investigación sobre problemas de polinización que causan bajas producciones en el ciruelo japonés.Proyecto de investigación CICYT AGL2006–13529–CO2–00 financiado por el Ministerio de Ciencia e Innovación–FEDE

Evaluation of the reproductive process as the cause for low fruit set in two japanese plum cultivars

The reproductive process has been evaluated to ascertain the causes of low fruit set in two Japanese plum cultivars, ‘Sweet August’ and ‘Rubirosa’. In order to evaluate pollen germination, pollen-pistil incompatibility and ovule viability, different experiments were performed in orchard conditions. Pollen viability was determined by assessing the percentage of in vitro pollen germination. The self-(in)compatibility of both cultivars and their cross-compatibility with ‘Champion’ were examined by self- and cross-pollination experiments followed by observation of pollen tube growth under the microscope. Likewise, the S-RNase alleles of each cultivar were determined by PCR amplification of the S-RNase gene. Finally, ovule development was examined under the microscope. While ‘Sweet August’ behaved as self-incompatible, ‘Rubirosa’ appears to be self-compatible. The observation of pollen germination and pollen tube growth in the different crosses analysed showed that fruit set was not limited by the absence of compatible pollen. However, a high percentage of degenerated ovules were observed in flowers from both cultivars. This premature ovule degeneration is likely to be the cause of low fruit set in both cultivars.Peer ReviewedsterilityS-allelefruit dropPrunus salicina Lindlovule degenerationpollinationPublishe

S-genotyping in Japanese plum by PCR and capillary gel electrophoresis detection

In this work a PCR S-genotyping method using capillary electrophoresis detection was assayed in Japanese plum. Sweet cherry primers designed for S-RNase and SFB intron length polymorphism detection by capillary electrophoresis were evaluated in Japanese plum cultivars. Amplification of both genes was successful and amplified sizes were correlated with Japanese plum S-alleles. The S-RNase genotype of 58 Japanese plum type cultivars previously determined by other methods was confirmed using this technology and the SFB alleles of these cultivars were also determined. Allele sizes of both genes are reported for 13 different S-alleles found in Japanese plum and will allow efficient S-genotype characterization in this species.Peer Reviewedgamenophytic self-incompatibilityS-RNaseSFBS-allelePrunus salicina LindlPublishe

S-Locus Genotyping in Japanese Plum by High Throughput Sequencing Using a Synthetic S-Loci Reference Sequence

Self-incompatibility in Prunus species is governed by a single locus consisting of two highly multi-allelic and tightly linked genes, one coding for an F-box protein—i.e., SFB in Prunus- controlling the pollen specificity and one coding for an S-RNase gene controlling the pistil specificity. Genotyping the allelic combination in a fruit tree species is an essential procedure both for cross-based breeding and for establishing pollination requirements. Gel-based PCR techniques using primer pairs designed from conserved regions and spanning polymorphic intronic regions are traditionally used for this task. However, with the great advance of massive sequencing techniques and the lowering of sequencing costs, new genotyping-by-sequencing procedures are emerging. The alignment of resequenced individuals to reference genomes, commonly used for polymorphism detection, yields little or no coverage in the S-locus region due to high polymorphism between different alleles within the same species, and cannot be used for this purpose. Using the available sequences of Japanese plum S-loci concatenated in a rosary-like structure as synthetic reference sequence, we describe a procedure to accurately genotype resequenced individuals that allowed the analysis of the S-genotype in 88 Japanese plum cultivars, 74 of them are reported for the first time. In addition to unraveling two new S-alleles from published reference genomes, we identified at least two S-alleles in 74 cultivars. According to their S-allele composition, they were assigned to 22 incompatibility groups, including nine new incompatibility groups reported here for the first time (XXVII-XXXV).Publishe

Simple Sequence Repeat (SSR)-Based Genetic Diversity in Interspecific Plumcot-Type (Prunus salicina × Prunus armeniaca) Hybrids

The main objective of many fruit-breeding programs around the world is the release of new cultivars from interspecific hybridizations between species of the Prunus genus. Plum × apricot (Prunus salicina Lindl. × Prunus armeniaca L.) are the most widespread interspecific hybrids, which include plumcots, pluots, and apriums. In this work, 115 accessions of interspecific hybrids from different origins and 27 reference genotypes of apricot and other diploid plum species were analyzed using eight simple sequence repeat (SSR) markers to assess the population structure and current genetic diversity. A total of 149 alleles were obtained, with an average of 19 alleles per locus. The overall polymorphic information content (PIC) mean value of SSR markers was 0.81, indicating a high degree of polymorphism of the SSR. The genetic analysis revealed 141 unique genotypes and two synonyms. The unweighted pair group method with arithmetic averages (UPGMA) dendrogram and the population structure with five groups inferred through the discriminant analysis of principal components (DAPC) revealed a clear genetic differentiation between apricot genotypes and the rest of the accessions since the interspecific hybrids clustered with the Japanese plum genotypes. Repeated backcrosses between interspecific hybrids with plum genotypes could be the cause of the higher genetic proximity of the hybrids with respect to plum than with apricot genotypes. This corresponds to the fruit morphology and agronomic behavior observed in most interspecific hybrids, which also resemble plums more than apricots.Publishe