Potential of genetic resistance of new table grape hybrids to fungal pathogens

Downy mildew (Plasmopara viticola) and powdery mildew (Erysiphe necator) are the most common and economically significant fungal diseases in vineyards. The task of this work is to study the genotypes of new promising hybrid forms of table grapes for the presence of resistance genes to downy mildew (Rpv10 and Rpv3) and powdery mildew (Ren9) using DNA-markers. The study was carried out on table grape hybrids under the working names Agat dubovskiy, Akelo, Arabella, Artek, Dubovskiy rozovyi, Gamlet, Ispolin, Kishmish dubovskiy, Kurazh, Pestryi, Valensiya and registered variety Liviya. The studied genes were analyzed using markers UDV305 and UDV737 (Rpv3), GF09-46 (Rpv10), CenGen6 (Ren9). The following cultivars were used as reference genotypes: Saperavi severnyi (carries Rpv10 gene) and Regent (Rpv3 and Ren9). It was established that Rpv3 gene is carried by hybrids Kishmish dubovskiy, Agat dubovskiy, Kurazh, Valensiya, Akelo, Gamlet, Dubovskiy rozovyi, Pestryi. Ren9 gene was found in Artek, Agat dubovskiy, Kurazh, Ispolin, Valensiya, Arabella, Gamlet, Dubovskiy rozovyi, Pestryi. The Rpv10 gene was not detected in any of the analyzed grapevine samples. genotypes Agat dubovskiy, Kurazh, Gamlet, Dubovskiy rozovyi, Pestryi, Valensiya carry Rpv3 and Ren9 genes simultaneously. These grapevines have an elegant bunch and large berries that are attractive to consumers.


Introduction
The development of DNA-marking methods has fundamentally changed the approaches to the classification and certification of varieties, the assessment of genetic diversity, mapping and identification of genes, genetic monitoring in genetics and breeding of cultivated plants. One of the areas of application of DNA-marker technologies is marker assisted selection (МАS), which allows identification and selection of genotypes carrying target genes, by passing phenotypic assessment, based only on DNA analysis data. DNA-markers are effectively used when working with grape genetic resources to study genetic diversity and biological potential, including resistance to the most harmful organisms.
DNA-markers identification of pathogen resistance genes in new breeding forms and grape varieties makes it possible to determine their adaptive potential to biotic stress factors. Downy mildew, caused by Plasmopara viticola, and powdery mildew, caused by Erysiphe necator, are the most common and economically significant fungal diseases of the vine. The determination of resistance genes in grapevines to these pathogens is carried out mainly in all world scientific centers [1][2][3][4][5][6][7][8][9]. Identification of genes for resistance to downy mildew and powdery mildew in domestic material with the aim of creating stable ampelocenoses, preserving the harvest and extending the productive life of grape plantations is an urgent task.
For fresh consumption, people use table grapes. The requirements for modern grape varieties for fresh consumption concern both the kind of the product (the size of the bunch, the size and shape of the berries, the color of the skin of the berries), its taste characteristics, as well as the transportability and suitability for storing the harvest, the processability of plantings and resistance to biotic and abiotic stressors. The resistance of the variety to pathogens allows to reduce chemical treatments during the growing process, which makes it possible to obtain environmentally safe products.
The task of this work was to study the genotypes of new promising table grapes hybrid forms for the presence of downy mildew (Rpv10 and Rpv3) and powdery mildew (Ren9) resistance genes using DNA markers.
Molecular genetic studies were carried out by the method of polymerase chain reaction (PCR) with the analysis of the results by the method of fragment analysis on an automatic genetic analyzer. DNA was extracted from young leaves of the apical part of the shoots by CTAB method [13].
PCR was carried out in a 20 μL reaction mixture containing nearly 50 ng of genomic DNA, 1.5 units of Tag polymerase, 1x buffer for Tag polymerase with ammonium sulfate and magnesium, 2 mM MgCl2, 0.2 mM each dNTP (deoxynucleotide triphosphates) (SibEnzyme-M, Moscow) and 200 μM of each primer (Syntol, Moscow) using a BioRad device (USA).
For the detection of Rpv10 gene, the GF09-46 marker was used [15], target PCR fragment indicating the presence of resistance allele did not found in any analyzed genotype (Table).
In the genotype of control variety Livia, with which the studied new grapevine hybrid forms are compared for complex of economically valuable traits, none of the analyzed resistance loci was found.

Conclusion
As a result of the DNA-marker analysis, it was found that Rpv3 gene is present in grapevine hybrids Kishmish dubovskiy, Agat dubovskiy, Kurazh, Valensiya, Akelo, Gamlet, Dubovskiy rozovyi, Pestryi (table). The powdery mildew resistance gene Ren9 was found in the following forms: Artek, Agat dubovskiy, Kurazh, Ispolin, Valensiya, Arabella, Gamlet, Dubovskiy rozovyi, Pestryi. The Rpv10 gene was not detected in any of the analyzed samples. Of the new hybrid forms of table grapes, the Rpv3 and Ren9 genes simultaneously carry the genotypes Agat dubovskiy, Kurazh, Gamlet, Dubovskiy rozovyi, Pestryi, Valensiya. These grapevine forms have beautiful, attractive bunch for the consumer and large berry ( fig.). They are promising for further study and for use in following breeding.