Search

CRISPR/Cas technology of genome editing is a powerful tool for making targeted changes in the DNA of various organisms, including plants. The choice of the precise nucleotide sequence (protospacer) in the gene to be edited is important in the design of guide RNA, which can be carried out by specialized software. We review and compare all the known on-line and off-line resources for guide RNA design, with special attention paid to tools capable of searching for off-target edits sites in plant genomes. The use of Cas12a may be preferable to Cas9. Techniques allowing C→T and G→A base editing without DNA cleavage are discussed along with the basic requirements for the design of effective and highly specific guide RNAs. Ways for improving guide RNA design software are presented. We also discuss the lesser risks of off-target editing in plant genomes as opposed to animal genomes. Examples of edited plant genomes including those that do not lead to the creation of transgenic plants are reviewed.

Amaranthus retroflexus was transformed with the pph6 gene encoding the synthesis of a metal-binding peptide, which on average increased plant resistance to Cd and Ni by 15%, the accumulation of heavy metals in plants increased by an average of 25%.

In order to investigate the effect of the RapA1 protein on growth-stimulating activity, biofilm formation, and root colonization by rhizobia, strains Rhizobium leguminosarum Pvu5, VSy12, Thy2, TPr4 and R. galegae 0702 were transformed with the pJN105TurboRapA1GFP vector, providing for constitutive expression of the rapA1 and gfp genes. Constitutive expression of the rapA1 gene was shown to result in more efficient biofilm formation on inert surfaces and to promote microcolony formation on plant root surfaces. Analysis of auxin production by wild and transformed strains revealed decreased auxin synthesis in all transformed strains, except for R. leguminosarum Thy2. Growth-stimulating activity of the wild and transformed strains did not correlate with the amount of auxins produced in vitro.

CRISPR/Cas technology of genome editing is a powerful tool for making targeted changes in the DNA of various organisms, including plants. The choice of the precise nucleotide sequence (protospacer) in the gene to be edited is important in the design of guide RNA, which can be carried out by specialized software. We review and compare all the known on-line and off-line resources for guide RNA design, with special attention paid to tools capable of searching for off-target edits sites in plant genomes. The use of Cas12a may be preferable to Cas9. Techniques allowing C→T and G→A base editing without DNA cleavage are discussed along with the basic requirements for the design of effective and highly specific guide RNAs. Ways for improving guide RNA design software are presented. We also discuss the lesser risks of off-target editing in plant genomes as opposed to animal genomes. Examples of edited plant genomes including those that do not lead to the creation of transgenic plants are reviewed.

Bacterial adhesins, the proteins responsible for attachment of plant growth-promoting rhizobacteria to plant roots, are involved in formation of stable associative symbioses. In the present work enhanced expression of the rapA1 adhesin gene in Rhizobium leguminosarum PVu5 was shown to improve the efficiency of nodulation on bean roots inoculated with the modified strain. The rapA1 gene was cloned into the pJN105Turbo plasmid, this construct was used for transformation of R. leguminosarum PVu5, bean plants were inoculated by this transgenic strain, and efficiency of root nodule formation was determined. In the plants treated with rapA1-transgenic rhizobia, the number of root nodules was on average two times higher than in the plants inoculated with the original strain. Aggregation of R. leguminosarum was achieved when the rapA1 gene expression was enhanced either in rhizobia or in the co-cultured modified strain E. coli pJN105TurboRapA1.