Blue barley grain pigmentation results from anthocyanin accumulation in the aleurone layer. Anthocyanins are known for their beneficial effects on human health. The gene encoding the MYELOCYTOMATOSIS 2 (MYC2) transcription factor is potentially responsible for the blue coloration of the aleurone. In non-pigmented barley, a single nucleotide insertion in this gene causes a frameshift mutation with a premature stop codon. It was hypothesized that restoring the MYC2 reading frame could activate anthocyanin accumulation in the aleurone. Using a targeted mutagenesis approach in the present study, the reading frame of MYC2 was restored in the non-pigmented cultivar Golden Promise. Genetic constructs harboring cas9 and gRNA expression units were developed, pre-validated in protoplasts, and then functional MYC2 alleles were generated at the plant level via Agrobacterium-mediated transformation. Anthocyanin accumulation in the aleurone layer of grains from these mutants was confirmed through microscopy and chemical analysis. The expression of anthocyanin biosynthesis genes was analyzed, revealing that the restoration of MYC2 led to increased transcript levels of F3H and ANS genes. These results confirm the critical role of the MYC2 transcription factor in the blue aleurone trait and provide a biotechnological solution for enriching barley grain with anthocyanins.

Background: Globodera rostochiensis belongs to major potato pathogens with a sophisticated mechanism of interaction with roots of the host plants. Resistance of commercial varieties is commonly based on specific R genes introgressed from natural populations of related wild species and from native potato varieties grown in the Andean highlands. Investigation of molecular resistance mechanisms and screening the natural populations for novel R genes are important for both fundamental knowledge on plant pathogen interactions and breeding for durable resistance. Here we exploited the Solanum phureja accessions collected in South America with contrasting resistance to G. rostochiensis. Results: The infestation of S. phureja with G. rostochiensis juveniles resulted in wounding stress followed by activation of cell division and tissue regeneration processes. Unlike the susceptible S. phureja genotype, the resistant accession reacted by rapid induction of variety of stress response related genes. This chain of molecular events accompanies the hypersensitive response at the juveniles’ invasion sites and provides high-level resistance. Transcriptomic analysis also revealed considerable differences between the analyzed S. phureja genotypes and the reference genome. Conclusion: The molecular processes in plant roots associated with changes in gene expression patterns in response to G. rostochiensis infestation and establishment of either resistant or susceptible phenotypes are discussed. De novo transcriptome assembling is considered as an important tool for discovery of novel resistance traits in S. phureja accessions.

Wild potato germplasm serves as a natural pool of agronomically valuable traits for potato breeding, such as resistance to pathogens and abiotic stresses, quality, and consumer-oriented traits. The introgression of these traits into cultivated potato is hampered by the different kinds of incompatibility and linkages between desirable and undesirable features in hybrid progeny. The trait donor improvement via correction of negative characteristics prior to hybridization to domestic potato can be a solution to the linkage drag problem. The de novo domestication concept for developing new crops using gene editing technologies was previously proposed and performed for tomato and physalis. In this review, we collected information about donor properties of different wild potato species and developed a strategy for potato germplasm enhancement using the de novo domestication approach. The possible modifications of several candidate genes responsible for undesirable traits in wild potato, including high steroidal glycoalkaloid content, self-incompatibility, tuberization under short day conditions, and long stolons are proposed. The current challenges and future prospects of implementing the de novo domestication strategy for potato are discussed.

—In this study, we performed expression analysis of genes associated with cold-induced sweetening in potato tubers: vacuolar invertase (Pain-1), sucrose synthase (SUS4), and invertase inhibitor (InvInh2). Potato varieties Nikulinsky, Symfonia, and Nevsky were used. All three varieties were found to accumulate sugars at low temperatures; the maximum accumulation of reducing sugars was observed at 4°C. It was found that the expression pattern of genes associated with cold-induced sweetening differs depending on the variety and storage duration. The increased expression of vacuolar invertase and its inhibitor is more pronounced at the beginning of storage period, whereas the increased expression of sucrose synthase is more pronounced after 3 months of storage. At early storage periods, high expression of invertase and low expression of inhibitor is observed in the Dutch variety Symfonia, and vice versa in the Russian varieties Nikulinsky and Nevsky. The involvement of the studied genes in the process of cold-induced sweetening is discussed.

Gene-editing tools enable precise, targeted genome modifications, providing new approach for the rapid and sustainable improvement of tea plant (Camellia sinensis (L.) Kuntze). Developing such an approach is especially important due to the perennial nature and complex genetics of the tea plant, which make traditional breeding slow and inefficient. To validate a gene editing protocol in the elite local tea cultivar Kolkhida three candidate genes were selected. Two guide RNAs (gRNAs) were designed for each gene, and corresponding constructs for targeted genome modification in tea were generated. Successful modifications of the target sequences in cv. Kolkhida tea protoplasts were achieved for all three target genes. The high mutagenic efficiency of the selected gRNAs was observed for two out of three genes, including induction of precise deletions between target motifs. gRNAs were delivered in protoplasts via co-transfection technique, and combined gRNA activity was observed when transfection efficiency exceeded 28%. The genome modification method for tea protoplasts established in this study can serve as a screening protocol to evaluate the in vivo efficiency of different genome editing approaches in the tea plant.