FROM THE EDITORIAL BOARD
METHODOLOGICAL ARTICLES (METHODOLOGY, METHODS, PROTOCOLS)
VIR is creating a collection of nomenclature standards for cultivars of different crops in line with the provisions of the International Code of Nomenclature for Cultivated Plants (ICNCP). The nomenclatural standards are designed in the form of a herbarium sample in cooperation with the authors of cultivars, who select plant material for each cultivar in the experimental field of their organization and forward it to the scientific herbarium together with the accompanying documents. Registering of herbarium specimen in the scientific herbarium is followed by the publication of nomenclatural standards. The article presents a new VIR genebank strategy for registering plant material of domestic cultivars received from breeders, which includes the design of nomenclatural standards; the development of a genetic passport of a cultivar using DNA samples from the plant delivered to the scientific herbarium; and application of biotechnological methods for keeping explants (isolated from the acquired plant material as buds or meristems) alive. The proposed integrated approach makes it possible to document cultivars not only with the help of nomenclatural standards, but also with the use of molecular-genetic methods, as well as to save genotyped specimens, transferred to herbarium, in in vitro and in cryo collections. This approach is applicable to any vegetatively propagated crop. The article summarizes the first results of our proposed strategy using modern domestic potato cultivars as an example. Collection, transfer and preparation of plant material for nomenclatural standards of potato varieties are specific; this article explains our protocol for this work. The implementation of this strategy in 2018-2020 led to the designation of nomenclatural standards for 66 Russian potato varieties and elaboration of their genetic passports, which are presented in a series of articles included into issues Nos.3 and 4, Volume 3, of the journal “Plant Biotechnology and Breeding”, containing an overview of methodological approaches to registration of modern domestic cultivars in the VIR collection. DNA samples obtained from the plant material provided by the cultivar authors for nomenclatural standard designation will continue to be used as a source of information on the genetic potential of a cultivar through the expansion of the list of DNA markers and the application of new genetic technologies.
In the present paper, the potato cultivars bred at the Leningrad Research Institute for Agriculture “Belogorka”, were taken as an example for demonstrating the results of elaboration of methodological approaches that are currently developed at the N.I. Vavilov Institute of Plant Genetic Resources (VIR) for the preparing of nomenclatural standards and their genotyping. In 2018, joint research of VIR scientists and breeders from the Leningrad Research Institute for Agriculture “Belogorka” began in the field of preparing nomenclatural standards for potato cultivars bred at this institute. Nomenclatural standards were prepared according to the ‘International Code of Nomenclature for Cultivated Plants’. Plant material for herbarium specimens was collected in the experimental field of the “Belogorka” Institute in 2018 by cultivar authors and handed over to the VIR Herbarium of cultivated plants, their wild relatives and weeds (WIR). The plant material included stems with inflorescences and later - tubers of 21 cultivars which were bred at the “Belogorka” Institute. Two precultivars undergoing State variety testing and three breeding clones were also included in this study. Just before herbarium preparation, the obtained plant material was photographed, plant morphological characters described, and the results compared with the description given in such official documents as the “Cultivar Questionnaireˮ and “Description of selection achievementˮ. The nomenclatural standards of 21 cultivars registered in the VIR Herbarium Database and transferred for conservation to the VIR herbarium, are published in this paper. Before herbarium preparation, the plant material was sampled for DNA extraction and subsequent genotyping and molecular screening. The genetic passports include information about the polymorphism of 10 chromosome-specific microsatellite loci, as well as the data on the presence/absence of diagnostic fragments of 12 markers of the 11 R-genes conferring resistance to diseases and pests, and for some cultivars – the information about their cytoplasm type. These genetic passports are valuable not only because different types of DNA markers were used in their preparing (SSR, SCAR and CAPS markers of the R genes; markers specific to different loci of the nuclear and organelle genomes), but first of all because of the material itself, as the DNA samples were isolated from the plants with the assigned status of nomenclatural standard for each particular cultivar. Based on the genetic passports data, trueness to type of the “Belogorka” cultivar samples obtained from various sources was verified.
Breeding of potato cultivars adapted to the climatic conditions of the Middle Volga region and resistant to various harmful organisms has always been an urgent task for breeders of the Tatarstan Republic. In the present paper, the potato cultivars bred at the Tatar Research Institute of Agriculture – Kazan Scientific Center of the Russian Academy of Sciences, were taken as an example for demonstrating the results of application and elaboration of methodological approaches that are currently developed at the N.I. Vavilov Institute of Plant Genetic Resources (VIR) for the preparing of nomenclatural standards and their genotyping. At the initiative of the VIR researchers in collaboration with the authors of potato cultivars E.A. Gimaeva and Z. Stashevski plant material was collected and nomenclatural standards for potato cultivars bred at the Kazan Scientific Center of the Russian Academy of Sciences were prepared. Four nomenclatural standards for Tatarstan potato cultivars (‘Zumba’, ‘Kortni’, ‘Reggi’, ‘Samba’) were prepared in accordance with the provisions of the International Code of Nomenclature For Cultivated Plants, registered in the VIR Herbarium Database, and transferred to the VIR herbarium type collection. Besides, voucher specimens of cv. ‘Tango’ and breeding clone ‘Sal’sa’, which is undergoing the State Variety Test at present, were produced. Genetic passports of Tatarstan varieties (‘Zumba’, ‘Kortni’, ‘Reggi’, ‘Samba’) and one breeding clone (‘Sal’sa’)* were developed using DNA samples from plant material transferred by the authors of the cultivars to the VIR herbarium. The genetic passports include information of the polymorphism of eight chromosome-specific microsatellites, data on the presence/ absence of the diagnostic fragments of 15 markers of the 11 R-genes conferring resistance to various harmful organisms and the information about cytoplasm types. A comparison of the data from genetic passports developed using DNA samples from nomenclatural standard specimens with the results of genotyping of 22 samples of Tatarstan cultivars obtained from different sources (e.g., samples undergoing ecogeographic tests within the framework of the Comprehensive Research Plan of the subprogram “Development of potato breeding and seed production in the Russian Federation”; as well as accessions from the VIR in vitro collection) made it possible to assess the authenticity and uniformity of the studied plant material.
ORIGINAL EXPERIMENTAL ARTICLES
Cryopreservation of shoot-tips (apexes) excised from in vitro plants is used for long-term preservation of potato cultivars. The most widely used method for creating potato cryo-collections is droplet-vitrification of shoot-tips which, together with its numerous modifications, is widely used in genebanks. A modified protocol of droplet vitrification method is used at VIR for cryopreservation of shoot tips from in vitro potato plants. This paper presents the results of cryopreservation of modern cultivars which were released by seven Russian breeding centers. In vitro clones used in the cryopreservation experiments were genetically identical to the cultivars’ nomenclatural standards and herbarium vouchers.The frequency of post-thaw regeneration in control experiments after short-term cryopreservation varied from 23.3 to 53.3%, depending on the genotype. Five out of 16 accessions (‘Varâg’, ‘Gusar’, ‘Evpatij’, ‘Solnečnyj’, ‘Tango’)* had low post-cryogenic regenerative capacity from 20 to 30%; the regeneration rate exceeded 30% in 11 accessions, and 8 cultivars (‘Grand’, ‘Zlatka’, ‘Lina’, ‘Safo’, ‘Siverskij’, ‘Signal’, ‘Utro’, ‘Ûna’) and ‘Аlyj Parus’ breeding clone had regeneration rate above 40%. The regeneration rate in the studied subset was genotype independent according to the ANOVA results (p=0.711). Viability and regeneration rate were significantly correlated (r=0.86). As a result of the experiments, explants of 14 modern cultivars and two breeding clones with the known post-thaw regeneration rate were successfully cryopreserved in the VIR cryobank. Four cultivars (‘Grand’, ‘Gusar’, ‘Signal’, ‘Utro’) were monitored for their regeneration capacity after the long-term (seven months) preservation in the VIR cryobank. On an average, these four cultivars demonstrated a post-thaw regeneration capacity of 41.8%. It can be concluded that the use of the modified method of droplet vitrification is relevant for increasing the VIR potato cryo-collection.
ISSN 2658-6258 (Online)