Delivery of CRISPR/Cas Components into Higher Plant Cells for Genome Editing

Russian Journal of Plant Physiology - CRISPR/Cas genome editing of plants is realized in three basic variants, including knockout mutations as indels, insertion of alien DNA fragments, and base...     

Protective Effect of Arachidonic Acid during Viral Infection: Synthesis of New Proteins by in vitroPotato Plants

The mechanisms of the protective, immunostimulating effects of arachidonic acid (AA) were studied, and its efficiency in the induction of defense reactions in the moderately virus-resistant potato cultivar Nevskii (Solanum tuberosumL.) was determined. Virus-free in vitropotato plants treated with AA and inoculated with phytopathogenic viruses were used as a model. The data on the X virus accumulation obtained by the enzyme-linked immunosorbent assay confirmed the immunizing effect of AA; the optimum concentration of the compound was 10^–8M. The antiviral effect of AA was maintained in infected in vitropotato plants for at least two or three weeks. The electrophoretic analysis of leaf proteins revealed a 33-kD polypeptide induced by the potato virus Y. Two weeks after inoculation with virus X, a 40-kD protein was identified in potato plants pretreated with AA. In addition, the relative content of the two groups of proteins consisting of two or three components with mol wts about 50 kD and above70 kD changed both upon viral infection and pretreatment with AA. Only small changes in the isozyme patterns of peroxidase in potato plants were observed during the development of systemic acquired resistance; they were manifested in some treatments in the band intensities. The existence of the alternative pathways of systemic acquired resistance in potato plants specifically activated by viral infection and AA was suggested.     

Hormonal status of tobacco variety Samsun NN exposed to synthetic coenzyme Q10 (ubiquinone 50) and TMV infection

Hormonal system status has been analyzed in leaf disks of hypersensitive tobacco Nicotiana tabacum L. variety Samsun NN during the development of resistance to tobacco mosaic virus (TMV) induced by synthetic coenzyme Q₁₀ (ubiquinone 50). The absolute and relative content of abscisic acid (ABA), indoleacetic acid (IAA), and cytokinins (CKs) was determined after the exposure of leaves to Q₁₀ solution and the subsequent TMV infection. In plants not treated with Q₁₀, CK content increased about 2.5 times 1 day after TMV infection, while a significant increase in the ABA level and a decrease in the IAA level were observed only after 2 days. In the dynamics, Q₁₀ treatment had a protective antiviral effect, significantly decreased the ABA level, and increased the IAA level in sensitized plants compared to nonsensitized ones.     

RAPD-PCR analysis of the variability of spring common wheat cultivar genomes and their androclinal double haploid form

Based on RAPD-PCR analysis with 15 primers including those homologous to particular loci, the level of genetic polymorphism in the collection of spring common wheat genotypes and their androclinal double haploid (ADH) lines was determined. Intraspecific polymorphism of the original wheat forms was 20%. By the absence of polymorphism in the molecular patterns of ADH lines and original forms, it was shown that, under the conditions used, no genetic changes in the genomic DNA of spring common wheat in anther culture occurred. Thus, the technology of direct in vitro androgenesis does not lead to genome rearrangements and may be used for rapid production of pure lines of such a complex allopolyploid as spring common wheat.     

Random Priming PCR Strategies for Identification of Multilocus DNA Polymorphism in Eukaryotes

A historical review of the advent and improvement of the methods for detecting multilocus DNA polymorphism that do not require preliminary knowledge of the individual gene and complete genome sequences of eukaryotes is presented. The first group of these methods includes approaches based on the use of primers with arbitrary sequence (random priming). Another group of methods to detect DNA polymorphism is based on the use of primers that consist of short repetitive sequences having anchor nucleotides at the 5'- or 3'-ends that position the annealing sites of these primers (microsatellite priming). Another approach for revealing polymorphism that does not require knowledge of the DNA sequence is based on cleavage of total DNA by a combination of restriction endonucleases (random cleavage) accompanied by PCR amplification. Considerable attention is paid to the opportunities of using these approaches to detect DNA polymorphism in the form of converting the obtained data to digital format and creation of integrative databases for all organisms, regardless of the methods used.     

Influence of arachidonic acid and phytoviruses on the hormonal system of potato plants grown in vitro

Changes in the phytohormonal system during the development of antivirus resistance was investigated. This resistance was induced by arachidonic acid (AA) in potato plants (Solanum tuberosum L.) grown in vitro. The plants in tubes were treated with AA 7 days before infection with tobacco mosaic virus or potato X virus. These plants were used for ABA, IAA, and cytokinin quantification. As a rule, phytoviruses increased ABA and cytokinin levels and decreased that of IAA in both immunized and unimmunized plants. AA significantly decreased the phytovirus content in potato plants at least during 3 weeks. In general, AA immunization acted as a limiting factor for increasing the ABA and cytokinin concentrations during virus pathogenesis in experimental plants. However, no regularity in the changes of the IAA concentration was noted. The data obtained showed that AA-induced changes in the hormonal system could be an important factor in the mechanism of plant resistance against phytopatogenic viruses.     

Design of Guide RNA for CRISPR/Cas Plant Genome Editing

Molecular Biology - 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...     

Protein composition of tobacco leaves with antiviral resistance induced by activators of defense responses and TMV infection

In tobacco (Nicotiana tabacum L.) plants of hypersensitive cv. Samsun NN, a capability of necrosis lesion formation and protein patterns were studied after induction of antiviral resistance by defense responses activators (DRA) (arachidonic acid, ubiquinone 50, and vitamin E) and by infection with tobacco mosaic virus (TMV). DRA and TMV improved both local and systemic leaf resistance to TMV. Native protein electrophoresis demonstrated differences in the composition of leaf proteins extracted under acidic and alkaline conditions. SDS-PAGE revealed proteins accumulated during the development of systemic antiviral resistance after lower leaf treatments with DRA and of local resistance induced by pretreatment with TMV. It was shown that various DRA affected protein patterns similarly, whereas TMV infection resulted in other changes. It is supposed that different pathways function in tobacco plants during induction of systemic resistance by DRA and TMV infection.     

Effect of ubiquinone 50 and viral infection on phytohemagglutinin activity in development of induced resistance in tobacco plants

The effects of ubiquinone 50 (synthetic coenzyme Q₁₀) and viral infection on phytohemagglutinin activity were studied in the tobacco (Nicotiana tabacum L.) cultivar Samsun NN, which carries the necrosis-formation locus and exhibits hypersensitivity to tobacco mosaic virus (TMV) and ubiquinone (Q₁₀). The treatment with Q₁₀ and inoculation with TMV were accompanied by an increase in phytohemagglutinin activity, indicating that lectins may be involved in the development of defense responses in plants.     

The Effect of Arachidonic Acid and Viral Infection on the Phytohemagglutinin Activity during the Development of Tobacco Acquired Resistance

The effects of arachidonic acid (AA) on the development of viral infection and the activity of phytohemagglutinins in Nicotiana tabacum L. plants were studied. Cv. Samsun NN was used, which displayed a genotypically determined hypersensitive response to tobacco mosaic virus (TMV) infection. When tobacco leaf disks were treated with 10^–9 to –10^–7 M AA, viral reproduction was suppressed by 90–100%. The AA concentration of 10^–8 M was optimal for the improvement of plant virus resistance. Tobacco leaves maintained virus resistance for at least two weeks. Both AA treatment and TMV inoculation were accompanied by an enhanced lectin activity, which may indicate the involvement of lectins in the development of plant defense responses. Lectin accumulation was observed in the intact plants developing systemic resistance and in the detached leaves characterized by local resistance.