Somatic Hybrids Research Articles

Mapping of Major QTLs Conferring Higher Branching Numbers in Backcross Introgression Lines of Sinapis alba + Brassica juncea Somatic Hybrids

Mapping of Major QTLs Conferring Higher Branching Numbers in Backcross Introgression Lines of Sinapis alba + Brassica juncea Somatic Hybrids

Interspecific hybridization and cell engineering of lettuce (Lactuca L.)

Lactuca sativa L. is a leafy vegetable crop of the Asteraceae family, widely cultivated throughout the world. The main breeding trends for lettuce include higher yields, better taste quality, earliness, and resistance to abiotic and biotic stressors. Some wild Lactuca spp. have actively been employed by lettuce breeders as donors of resistance to various diseases. Conventional and biotechnological breeding methods are both currently used to develop new lettuce cultivars. This is an overview of the main advances in the production of interspecific Lactuca hybrids, including the use of cell and tissue culture techniques, and genetic engineering. Studying artificial hybridization and natural populations makes it possible to identify evolutionary relationships among various Lactuca spp. Somatic hybridization is an overlooked but promising technology in Lactuca breeding: it allows a breeder to obtain a wider range of variations, and is beyond the strict control by GMO laws. This technique faces problems associated with complicated protoplast regeneration and the loss of reproductive ability in hybrids. Genome-editing methods are more effective and better controllable, but society is still wary of any interference with the plant genome and legally regulates the sale of GM products as food. Thus, researchers are challenged with the task to improve these techniques.

In vivo tetraploid induction of mono-embryonic citrus genotypes by colchicine treatment

Polyploid holds great significance in citrus breeding, to produce dwarf tetraploid rootstocks with wide adaptability and triploid scion varieties bearing seedless fruits. In citrus, tetraploids have been generated through somatic hybridization and autonomous chromosome doubling of nucellar embryos. However, most tetraploids were recovered from poly-embryonic (apomictic) citrus, while tetraploids of mono-embryonic (non-apomictic) citrus were seldomly reported. Herein, we developed an in vivo tetraploid induction method by colchicine treatment, which is especially high-efficient in mono-embryonic citrus. The epicotyl of citrus seedling was decapitated and incubated in dark for different time durations (0–15 d) to induce callus formation on the epicotyl wound. The callus on wound was then immersed in different concentrations (0.0125 %-0.100 %) of colchicine for 2 h, enabling regeneration of tetraploid shoots. We achieved high tetraploid induction rate (20.69 %) but low mixoploid induction rate (1.72 %), when 0.025 % colchicine was applied on wound cultured in dark for 9 d By using this method, 147 tetraploids were recovered from mono-embryonic mandarin, tangerine, sweet orange, pummelo and citron. Our method exhibited high tetraploid induction efficiency (14.28 %-20.69 %), low rate of mixoploid (1.72 %-5.30 %), short recovery period (45 d), and no need of tissue culture. The majority (75 %-100 %) of the recovered tetraploids maintained stable ploidy during subsequent growth, and the mono-embryonic nature of the tetraploids was identified by InDel marker analysis. The mono-embryonic tetraploids obtained herein enriched polyploid germplasm pool for citrus breeding.

Modern Technologies Provide New Opportunities for Somatic Hybridization in the Breeding of Woody Plants.

Advances in cell fusion technology have propelled breeding into the realm of somatic hybridization, enabling the transfer of genetic material independent of sexual reproduction. This has facilitated genome recombination both within and between species. Despite its use in plant breeding for over fifty years, somatic hybridization has been limited by cumbersome procedures, such as protoplast isolation, hybridized-cell selection and cultivation, and regeneration, particularly in woody perennial species that are difficult to regenerate. This review summarizes the development of somatic hybridization, explores the challenges and solutions associated with cell fusion technology in woody perennials, and outlines the process of protoplast regeneration. Recent advancements in genome editing and plant cell regeneration present new opportunities for applying somatic hybridization in breeding. We offer a perspective on integrating these emerging technologies to enhance somatic hybridization in woody perennial plants.

Wild Genetic Resource in Vegetable Improvement: Applications and Strategies

Domestication of feral types of the present-day cultivated types was in vogue in earlier days of farming followed by the selection and hybridization that led to many improved varieties. Expression of traits faded due to the development of homozygosity due to repeated interventions in the same set. Relying on wild relatives that are found to be the source of many target genes is found significant. While, this is not so easy, because of the barriers that reside in distant hybridization, consequences like failure in zygote and embryo formation, embryo death, linkage drags, and incompatibility within wild and cultivated groups, etc are need of concern. Strategies like bridge crosses, somatic hybridization, grafting, embryo rescue, etc., techniques are solely or in combination can combat these issues. Vegetables are the group of crops that are mainly cultivated for their high returns and quality aspects for the healthy food routine of the consumer. This review will spotlight the recent advances in vegetable improvement through their wild relatives and available strategies to overcome the barriers to the usage of wild relatives in vegetable improvement, meanwhile the conservation of these resources for the future utilization in vegetable improvement.

Genetic diversification of allohexaploid Brassica hybrids (AABBCC) using a fertile octoploid with excessive C genome set (AABBCCCC).

Using octoploid somatic hybrids with excessive C genome sets, AABBCCCC, a diverse allohexaploid, AABBCC, was produced by C genome reduction through subsequent crossing with various AABB cultivars. Even when somatic hybrids are produced, the plants that are produced are rarely in themselves an innovative crop. In this study, we used somatic hybrids of Brassica juncea (AABB) and B. oleracea (CC) as model cases for the genetic diversification of the somatic hybrids. One cell of 'Akaoba Takana' (B. juncea) and two cells of 'Snow Crown' (B. oleracea) were fused to create several somatic hybrids with excessive C genomes, AABBCCCC. Using AABBCCCC somatic hybrids as mother plants and crossing with 'Akaoba Takana', the AABBCC progenies were generated. When these AABBCC plants were self-fertilized, and flow cytometric (FCM) analysis was performed on the next generations, differences in the relative amount of genome size variation were observed, depending on the different AABBCCCC parents used for AABBCC creation. Further self-progeny was obtained for AABBCC plants with a theoretical allohexaploid DNA index by FCM. However, as the DNA indices of the progeny populations varied between plants used and aneuploid individuals still occurred in the progeny populations, it was difficult to say that the allohexaploid genome was fully stabilized. Next, to obtain genetic diversification of the allohexaploid, different cultivars of B. juncea were crossed with AABBCCCC, resulting in diverse AABBCC plants. Genetic diversity can be further expanded by crossbreeding plants with different AABBCC genome sets. Although genetic stability is necessary to ensure in the later generations, the results obtained in this study show that the use of somatic hybrids with excess genomes is an effective strategy for creating innovative crops.

Transcriptome analysis reveals genes associated with late blight resistance in potato

Late blight is a serious disease of potato worldwide. Our study aimed to unveil genes involved in late blight resistance in potato by RNA-seq analysis after artificial inoculation under controlled conditions. In this study, two potato somatic hybrids (P7 and Crd6) and three varieties such as Kufri Girdhari, Kufri Jyoti and Kufri Bahar (control) were used. Transcriptiome analysis revealed statistically significant (p < 0.05) differentially expressed genes (DEGs), which were analysed into up-regulated and down-regulated genes. Further, DEGs were functionally characterized by the Gene Ontology annotations and the Kyoto Encyclopedia of Genes and Genomes pathways. Overall, some of the up-regulated genes in resistant genotypes were disease resistance proteins such as CC-NBS-LRR resistance protein, ankyrin repeat family protein, cytochrome P450, leucine-rich repeat family protein/protein kinase family, and MYB transcription factor. Sequence diversity analysis based on 38 peptide sequences representing 18 genes showed distinct variation and the presence of three motifs in 15 amino acid sequences. Selected genes were also validated by real-time quantitative polymerase chain reaction analysis. Interestingly, gene expression markers were developed for late blight resistant genotypes. Our study elucidates genes involved in imparting late blight resistance in potato, which will be beneficial for its management strategies in the future.

Modern Breeding Strategies and Tools for Durable Late Blight Resistance in Potato.

Cultivated potato (Solanum tuberosum) is a major crop worldwide. It occupies the second place after cereals (corn, rice, and wheat). This important crop is threatened by the Oomycete Phytophthora infestans, the agent of late blight disease. This pathogen was first encountered during the Irish famine during the 1840s and is a reemerging threat to potatoes. It is mainly controlled chemically by using fungicides, but due to health and environmental concerns, the best alternative is resistance. When there is no disease, no treatment is required. In this study, we present a summary of the ongoing efforts concerning resistance breeding of potato against this devastating pathogen, P. infestans. This work begins with the search for and selection of resistance genes, whether they are from within or from outside the species. The genetic methods developed to date for gene mining, such as effectoromics and GWAS, provide researchers with the ability to identify genes of interest more efficiently. Once identified, these genes are cloned using molecular markers (MAS or QRL) and can then be introduced into different cultivars using somatic hybridization or recombinant DNA technology. More innovative technologies have been developed lately, such as gene editing using the CRISPR system or gene silencing, by exploiting iRNA strategies that have emerged as promising tools for managing Phytophthora infestans, which can be employed. Also, gene pyramiding or gene stacking, which involves the accumulation of two or more R genes on the same individual plant, is an innovative method that has yielded many promising results. All these advances related to the development of molecular techniques for obtaining new potato cultivars resistant to P. infestans can contribute not only to reducing losses in agriculture but especially to ensuring food security and safety.

IbNF-YA1 is a key factor in the storage root development of sweet potato.

As a major worldwide root crop, the mechanism underlying storage root yield formation has always been a hot topic in sweet potato [Ipomoea batatas (L.) Lam.]. Previously, we conducted the transcriptome database of differentially expressed genes between the cultivated sweet potato cultivar "Xushu18," its diploid wild relative Ipomoea triloba without storage root, and their interspecific somatic hybrid XT1 with medium-sized storage root. We selected one of these candidate genes, IbNF-YA1, for subsequent analysis. IbNF-YA1 encodes a nuclear transcription factor Y subunit alpha (NF-YA) gene, which is significantly induced by the natural auxin indole-3-acetic acid (IAA). The storage root yield of the IbNF-YA1 overexpression (OE) plant decreased by 29.15-40.22% compared with the wild type, while that of the RNAi plant increased by 10.16-21.58%. Additionally, IAA content increased significantly in OE plants. Conversely, the content of IAA decreased significantly in RNAi plants. Furthermore, real-time quantitative reverse transcription-PCR (qRT-PCR) analysis demonstrated that the expressions of the key genes IbYUCCA2, IbYUCCA4, and IbYUCCA8 in the IAA biosynthetic pathway were significantly changed intransgenic plants. The results indicated that IbNF-YA1 could directly target IbYUCCA4 and activate IbYUCCA4 transcription. The IAA content of IbYUCCA4 OE plants increased by 71.77-98.31%. Correspondingly, the storage root yield of the IbYUCCA4 OE plant decreased by 77.91-80.52%. These findings indicate that downregulating the IbNF-YA1 gene could improve the storage root yield in sweet potato.

An international breeding project using a wild potato relative Solanum commersonii resulted in two new frost-tolerant native potato cultivars for the Andes and the Altiplano.

This breeding project, initiated at the United States Potato Genebank (USPG) in collaboration with Peruvian partners Instituto Nacional de Innovacion Agraria (INIA), International Potato Center, Peru (CIP), and local farmers, sought to enhance cold hardiness and frost tolerance in native potato cultivars in Peru. The Andes and Altiplano are often affected by frost, which causes significant reduction in yield; creating varieties with superior resilience is a critical undertaking. The goal was to transfer outstanding non-acclimated cold tolerance and acclimation capacity found in wild potato species Solanum commersonii (cmm). Breeding families segregating for cold hardiness were created using (a) a somatic hybrid cmm + haploid Solanum tuberosum (tbr) (cv. Superior, US variety from Wisconsin) as male and (b) seven cultivars native to Peru of the species S. tuberosum sbsp. andigenum (adg) as females. All plant materials were part of the USPG germplasm collection. Sexual seeds of each family were sent to Peru for evaluations under the natural conditions of the Andean highlands and Altiplano. The plants were assessed for their response to frost, and genotypes showing exceptional tolerance were selected. Plants were also evaluated for good tuber traits and yield. Initial planting involving ~2,500 seedlings in five locations resulted in selecting 58 genotypes with exceptional frost tolerance, good recovery capacity after frost, and good tuber traits. Over the years, evaluations continued and were expanded to replicated field trials in the harsher conditions of the Altiplano (Puno). All trials confirmed consistency of frost tolerance over time and location, tuber quality, and yield. After 8 years, two advanced clones were considered for cultivar release because of their exceptional frost tolerance and superior field productivity that outyielded many of the established cultivars in the region. In November 2018, a new native cultivar named Wiñay, a Quechua word meaning "to grow" was released in Peru. In 2022, a second cultivar followed with the name Llapanchispaq (meaning "for all of us"). This project evidenced that a multinational and all-encompassing approach to deploy valuable genetic diversity can work and deliver effective results. This is even more significant when outcomes can promote food security and sustainability in very vulnerable regions of the world.

Enhancing Protoplast Isolation and Early Cell Division from Cannabis sativa Callus Cultures via Phenylpropanoid Inhibition.

De novo regeneration of Cannabis sativa L. (cannabis) using tissue culture techniques remains unreliable and infrequent. Conventional methods for the regeneration and transformation of cannabis have not achieved the reliability and replicability that need to be integrated into research and breeding programs. Protoplast systems are effective for gene expression studies and transformation and genome-editing technologies and open the possibility of somatic hybridization to create interspecific hybrids. To date, leaf-derived protoplasts have been isolated for transient gene expression studies, but protoplast-to-plant regeneration has not been reported. The present study aims to evaluate the efficacy of using a callus culture system as an abundant tissue source for protoplast isolation and lays the groundwork for a protoplast-to-plant regeneration system. Using hypocotyl-derived callus cultures, which are known to have relatively greater regenerative potential, the efficacy of protoplast isolation and initial cell division were assessed. In this study, the effect of 2-aminoindane-2-phosphonic acid (AIP), a competitive inhibitor of phenylalanine ammonia lyase (PAL), in callus culture media and the effect of subculture frequency on protoplast yield were assessed. This study found that inclusion of AIP at 1 mM resulted in a 334% increase in protoplast yield compared with AIP-free medium, representing the first known use of AIP in cannabis tissue culture. Inclusion of AIP led to a 28% decrease in total soluble phenolics and 52% decrease in tissue browning compared with the control medium. Lastly, a two-phase culture system for protoplast regeneration was tested. At a concentration of 2.0 × 105 protoplasts per mL, cell wall reconstitution and cell division were observed, providing one of the first know reports of cell division from cannabis protoplasts and setting the stage for the future development of a protoplast-to-plant regeneration system.

Somatic hybridization on different horticultural crops: A review

Somatic hybridization on different horticultural crops: A review

Potato: Breeding and Genomics

The commonly cultivated potato is a tetraploid and belongs to the species Solanum tuberosum, which includes two subspecies viz. ssp. tuberosum adapted to long days and ssp. Andigena adapted to short days. The conventional potato breeding programmes depend mainly on the identification of promising parental lines for making desired crosses, the creation of genetic variability through the crossing and subsequently the selection of desirable recombinants for further evaluation and vegetative propagation. During the last seven decades, Indian Council of Agricultural Research - Central Potato Research Institute (ICAR -CPRI) has developed and released as many as 69 improved varieties and one TPS population. These varieties cater to the need of the farmers across the country; and have superior agronomic attributes and resistance to different biotic and abiotic stresses. The somatic hybrids has been produced successfully by utilization of wild solanum species like S. etuberosum, S. cardiophyllam and S. pinnetisectum. Further, considerable work has been done in potato applying molecular biology and genomics approaches to address biotic and abiotic stress and quality traits.

Selection and Phenotyping for Drought Tolerance in Somatic Hybrids between Solanum tuberosum and Solanum bulbocastanum That Show Resistance to Late Blight, by Using a Semi-Automated Plant Phenotyping Platform

Drought stress is one of the most limiting abiotic stresses for plant growth and development. Potato (Solanum tuberosum L.), due to its shallow root system, is considered sensitive to drought. In potato breeding, the wild Solanum species may represent a good resource for disease and abiotic stress resistance genes, but their transfer is limited by sexual incompatibilities. Somatic hybrids (SH) between potato and the wild species Solanum bulbocastanum, sexually incompatible with potato, proved to be late-blight-resistant in laboratory and field assays. The aim of this study was to screen a series of somatic hybrids and derived backcrosses for drought stress tolerance. In vitro stress exposure (with 5% and 15% PEG 6000) allowed the selection of several tolerant genotypes in a short time. The eleven selected genotypes were tested by using a semi-automated plant phenotyping platform at the Biological Research Centre in Szeged, Hungary, where the plants’ biomass accumulation and photosynthesis under long-term drought conditions were monitored. The findings of this study affirm that the somatic hybrids between potato and S. bulbocastanum, along with their backcrosses, constitute valuable pre-breeding material. This is attributed to their possession of both late blight resistance and drought stress tolerance.

Physiological and Biochemical Evaluation of Salt Stress Tolerance in a Citrus Tetraploid Somatic Hybrid

Somatic hybridization has emerged as a valuable tool for developing novel genetic combinations in citrus breeding programs, including the creation of salt-tolerant rootstocks. In this study, the performance of a tetraploid somatic hybrid, obtained by fusing protoplasts derived from salt-tolerant Cleopatra mandarin (Citrus reshni hort. ex Tanaka) and salt-sensitive Carrizo citrange (Citrus sinensis L. Osbeck × Poncirus trifoliata L. Raf), was assessed under in vitro salt stress. Hybrid plants were characterized by leaf morphology, and ploidy level by flow cytometry and molecular markers. In vitro shoots were generated from the micropropagation of mature stem pieces of the somatic hybrid and its parents, and these were challenged by exposure to NaCl (0, 50, or 100 mM) supplemented to the media for three weeks to induce salt stress. The leaves of the somatic hybrid display intermediate morphology compared to the parental Cleopatra mandarin and Carrizo citrange rootstocks. All molecular markers successfully amplified DNA from the three cultivars; however, only 11 of 14 unequivocally confirmed somatic hybridity. The physiological and biochemical parameters, including chlorophyll content, lipid peroxidation, total phenolic compounds, antioxidants activity and proline content, were measured in the leaves. The somatic hybrid exhibited superior salt stress tolerance compared to the parent varieties, as evidenced by the reduced cellular membrane damage indicated by the lower levels of malondialdehyde and electrolyte leakage, particularly under 100 mM NaCl treatment. The somatic tetraploid hybrid also displayed higher total phenolic content than either parent, while Cleopatra mandarin exhibited the highest proline levels under 50 mm NaCl. These results demonstrate the enhanced salinity stress tolerance of the somatic hybrid compared to its parent lines, highlighting its potential as a valuable candidate for developing salt-tolerant citrus rootstocks.

A Comprehensive Review on Role of Plant Tissue Culture in Ornamental Crops: Cultivation Factors, Applications and Future Aspects

Ornamental plants are grown largely for their artistic value, floriculturists must prioritize the proliferation and improvement of quality traits, as well as the production of unique diversity. Micropropagation, clonal reliability and conservation are all crucial factors to consider. Application of in vitro techniques in ornamental plant such as in vitro embryo rescue, somatic hybridization, in vitro pollination and in vitro ploidy manipulation but to enhance, techniques like as embryo rescue and somatic hybridization are commonly employed. The creation of synthetic seed allows for season-independent seed producing and long-term seed preservation. Many factors influence ornamental plant tissue culture, including plant genotype, explants type, and the physical environment (light, temperature, humidity, and CO2), in addition to medium composition and growth regulators. We compiled and reviewed an overall update on cultivation factors, application procedures in ornamental plant tissue culture, in vitro plant enhancement approaches and future prospects in this study.

Protoplast Isolation and Fusion in Amomum Subulatum Roxb

Amomum subulatum Roxb. is an important spice and cash crop of Himalayan region of Indian sub-continent. An attempt has been made to isolate and fuse protoplast from the in-vitro grown seedling of Amomum subulatum Roxb. Off the various enzyme combination tried, the one containing the enzyme combination of 2% cellulase “Ozonuka” RS (Yakult Pharmaceutical Ind. Co. Ltd, Japan) 1% Macerozyme R-10 (Yakult Pharmaceutical Ind. Co. Ltd, Japan) and 1 % Hemi-cellulase (Sigma-Aldrich, USA) prepared in 10.93% D- Mannitol (Himedia, India) and 0.055% calcium chloride yielded protoplast of 2 x 104/g of leaf tissue. Protoplast fusion was achieved using 20% Polyethylene Gylcol (PEG 4000) but in small quantity added to the mixed protoplast. To develop a successful somatic hybrid of commercial importance further research is required.

Early selection of carrot somatic hybrids: a promising tool for species with high regenerative ability

BackgroundSince its discovery, somatic hybridization has been used to overcome the sexual barriers between cultivated and wild species. A combination of two somatic cells might provide a novel set of features, often of agronomical importance. Here, we report a successful approach for production and selection of interspecific somatic hybrid plants between cultivated and wild carrot using dual-labelling of protoplasts and an early selection of fused cells via micromanipulator. Both subspecies used in this study are characterised by a very high regenerative ability in protoplast cultures. Thus, a precise and effective method of hybrid selection is essential to assure the development and regeneration of much less numerous heterokaryons in the post-fusion cell mixture.ResultsElectrofusion parameters, such as alternating current and direct current, were optimised for an efficient alignment of protoplasts and reversible membrane breakdown followed by a cell fusion. Four hundred twenty-nine cells emitting green–red fluorescence, identified as hybrids, were obtained. Co-culture with donor-derived protoplasts in the alginate feeder layer system stimulated re-synthesis of the cell wall and promoted cell divisions of fusants. Somatic embryogenesis occurred in hybrid-derived microcalli cultures, followed by plant regeneration. Regenerated hybrids produced yellowish storage roots and leaves of an intermediate shape between cultivated and wild subspecies. The intron length polymorphism analysis revealed that 123 of 124 regenerated plants were hybrids.ConclusionsThe developed protocol for protoplast fusion and an early selection of hybrids may serve as an alternative to combining genomes and transferring nuclear or cytoplasmatic traits from wild Daucus species to cultivated carrot.

Identification of novel late blight resistance source in wild potato species and interspecific somatic hybrids, and their distinctness, uniformity and stability (DUS) characterization

Forty potato genotypes, including wild Solanum species and interspecific somatic hybrids were characterized for late blight resistance and distinctness, uniformity and stability (DUS) descriptors. Thirty-two genotypes (14 wild species + 18 somatic hybrids) were found highly resistant to late blight, whereas 4 were resistant and 1 was moderately resistant compared to the control varieties viz., Kufri Jyoti (susceptible), Kufri Bahar (highly susceptible), and Kufri Girdhari (highly resistant). All wild species were high resistant to late blight. Further, morphological characterization based on 51 DUS descriptors showed phenotypic variation in the genotypes studied. Thus, we identified late blight-resistant potato wild species and somatic hybrids, which could serve as a potential source for late blight resistance breeding.

Promotive effect of phytosulfokine - peptide growth factor - on protoplast cultures development in Fagopyrum tataricum (L.) Gaertn

BackgroundFagopyrum tataricum (Tartary buckwheat) is a valuable crop of great nutritional importance due to its high level of bioactive compounds. Excellent opportunities to obtain plants with the high level or the desired profile of valuable metabolites may be provided by in vitro cultures. Among known in vitro techniques, protoplast technology is an exciting tool for genetic manipulation to improve crop traits. In that context, protoplast fusion may be applied to generate hybrid cells between different species of Fagopyrum. To apply protoplast cultures to the aforementioned approaches in this research, we established the protoplast-to-plant system in Tartary buckwheat.ResultsIn this work, cellulase and pectinase activity enabled protoplast isolation from non-morphogenic and morphogenic callus (MC), reaching, on average, 2.3 × 106 protoplasts per g of fresh weight. However, to release protoplasts from hypocotyls, the key step was the application of driselase in the enzyme mixture. We showed that colony formation could be induced after protoplast embedding in agarose compared to the alginate matrix. Protoplasts cultured in a medium based on Kao and Michayluk supplemented with phytosulfokine (PSK) rebuilt cell walls, underwent repeated mitotic division, formed aggregates, which consequently led to callus formation. Plating efficiency, expressing the number of cell aggregate formed, in 10-day-old protoplast cultures varied from 14% for morphogenic callus to 30% for hypocotyls used as a protoplast source. However plant regeneration via somatic embryogenesis and organogenesis occurred only during the cultivation of MC-derived protoplasts.ConclusionsThis study demonstrated that the applied protoplast isolation approach facilitated the recovery of viable protoplasts. Moreover, the embedding of protoplasts in an agarose matrix and supplementation of a culture medium with PSK effectively stimulated cell division and further development of Tartary buckwheat protoplast cultures along with the plant regeneration. Together, these results provide the first evidence of developing a protoplast-to-plant system from the MC of Fagopyrum tataricum used as source material. These findings suggest that Tartary buckwheat’s protoplast cultures have potential implications for the species’ somatic hybridization and genetic improvement.