The activity of the pool of sgRNA molecules designed for different regions of potato coilin and phytoene desaturase genes was compared in vitro. Due to the presence of nucleotides unpaired with DNA, sgRNA is able not only to inhibit but also to stimulate the activity of the Cas9–sgRNA complex in vitro. Although the first six nucleotides located in the DNA substrate proximally to the PAM site at the 3' end are the binding sites for cas9, they had no significant effect on the activity of the Cas9–sgRNA complex.

Tomato (Solanum lycopersicum L.) is one of the most commercially essential vegetable crops cultivated worldwide. In addition to the nutritional value, tomato is an excellent model for studying climacteric fruits’ ripening processes. Despite this, the available natural pool of genes that allows expanding phenotypic diversity is limited, and the difficulties of crossing using classical selection methods when stacking traits increase proportionally with each additional feature. Modern methods of the genetic engineering of tomatoes have extensive potential applications, such as enhancing the expression of existing gene(s), integrating artificial and heterologous gene(s), pointing changes in target gene sequences while keeping allelic combinations characteristic of successful commercial varieties, and many others. However, it is necessary to understand the fundamental principles of the gene molecular regulation involved in tomato fruit ripening for its successful use in creating new varieties. Although the candidate genes mediate ripening have been identified, a complete picture of their relationship has yet to be formed. This review summarizes the latest (2017–2023) achievements related to studying the ripening processes of tomato fruits. This work attempts to systematize the results of various research articles and display the interaction pattern of genes regulating the process of tomato fruit ripening.

The tomato is a convenient object for studying reproductive processes, which has become a classic. Such complex processes as flowering and fruit setting require an understanding of the fundamental principles of molecular interaction, the structures of genes and proteins, the construction of signaling pathways for transcription regulation, including the synchronous actions of cis-regulatory elements (promoter and enhancer), trans-regulatory elements (transcription factors and regulatory RNAs), and transposable elements and epigenetic regulators (DNA methylation and acetylation, chromatin structure). Here, we discuss the current state of research on tomatoes (2017–2023) devoted to studying the function of genes that regulate flowering and signal regulation systems using genome-editing technologies, RNA interference gene silencing, and gene overexpression, including heterologous expression. Although the central candidate genes for these regulatory components have been identified, a complete picture of their relationship has yet to be formed. Therefore, this review summarizes the latest achievements related to studying the processes of flowering and fruit set. This work attempts to display the gene interaction scheme to better understand the events under consideration.

Various plant-derived promoters can be used to regulate ectopic gene expression in potato. In the present study, four promoters derived from the potato genome have been characterized by the expression of identical cassettes carrying the fusion with the reporter β-glucuronidase (gusA) gene. The strengths of StUbi, StGBSS, StPat, and StLhca3 promoters were compared with the conventional constitutive CaMV 35S promoter in various organs (leaves, stems, roots, and tubers) of greenhouse-grown plants. The final amount of gene product was determined at the post-transcriptional level using histochemical analysis, fluorometric measurements, and Western blot analysis. The promoter strength comparison demonstrated that the StUbi promoter generally provided a higher level of constitutive β-glucuronidase accumulation than the viral CaMV 35S promoter. Although the StLhca3 promoter was predominantly expressed in a green tissue-specific manner (leaves and stems) while StGBSS and StPat mainly provided tuber-specific activity, a “promoter leakage” was also found. However, the degree of unspecific activity depended on the particular transgenic line and tissue. According to fluorometric data, the functional activity of promoters in leaves could be arranged as follows: StLhca3 > StUbi > CaMV 35S > StPat > StGBSS (from highest to lowest). In tubers, the higher expression was detected in transgenic plants expressing StPat-gusA fusion construct, and the strength order was as follows: StPat > StGBSS > StUbi > CaMV 35S > StLhca3. The observed differences between expression patterns are discussed considering the benefits and limitations for the usage of each promoter to regulate the expression of genes in a particular potato tissue.