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Bioinformatic analysis of nucleotide sequences of the acetolactate synthase (als) gene of different members of the Poaceae family
2020
Кирієнко, А. В | Щербак, Н. Л | Симоненко, Ю. В
Purpose. Provide bioinformatic analysis and comparison of target regions of the acetolactate synthase (als) gene in several members of the Poaceae family and, on the basis of the obtained data, explore the possibility of creating a unified genetic construct for als gene editing using the CRISPR-Cas9 system. Methods. The als gene sequences of various members of the Poaceae family were obtained from the NCBI: Nucleotide database. For comparison, a fragment of the imi-2 gene of wheat of the soft line ‘TealIMI11A’ was used in two regions of the 367–390 and 1729–1749 nucleotide sequences. The Sequence Viewer 3.37.0 tool was used to assess the presence of nucleotide substitutions in the working sequence of the imi-2 gene. The dendrogram was built using the “Blast Tree” tool from the NCBI: Blast: Nucleotide resource. Results. A comparative analysis of the nucleotide sequences of seven different species was carried out: soft wheat (Triticum aestivum L.), common wild oat (Avena fatua L.), barley (Hordeum vulgare L.), Asian rice (Oryza sativa L.), maize (Zea mays L.), aleppo grass (Sorghum halepense Pers.) and Tausch’s goatgrass (Aegilops tauschii Coss.). The dendrogram is based on the gene sequence als, showed that all studied genotypes can be divided into two blocks: the first block included maize and aleppo grass, and the second block, a separate branch includes Asian rice and common wild oat, barley, soft wheat and Tausch’s goatgrass. 367–390 nucleotide sequences of soft wheat showed the highest 100% homology to Asian rice, Tausch’s goatgrass and common wild oat. The lowest homology was for maize and aleppo grass at 83.3%. Evaluation of the nucleotide sequence 1729–1749 showed no complete homology at the 100% level. It was the highest for barley and Tausch’s goatgrass – 95.2%, and the lowest for rice, maize and aleppo grass – 80.9% each. Conclusions. The analysis confirms a significant degree of homology of the als gene sequence for various species of the Poaceae family, which allows us to create a universal genetic vector. However, taking into account the high degree of sequence homology for species such as soft wheat, Tausch’s goatgrass, barley, Asian rice and common wild oat, it can be assumed that the corresponding genetic vector can be used with the greatest efficiency to alter the als gene of these genotypes.
显示更多 [+] 显示较少 [-]Bioinformatic analysis of maize gene encoding starch branching enzyme SBEIIb
2016
Сліщук, Г. І | Жернаков, Т. Ю | Волкова, Н. Е
Purpose. Investigation of maize ae1 gene polymorphism by bioinformatic methods. Methods. Global and local alignment of the nucleotide and amino acid sequences, in silico translation and transcription, translates modeling, primers design, phylogenetic analysis. Results. 255 nucleotide sequences of maize аe1 gene, 500 amino acid sequences of homology translates of maize ae1 gene (SBEIIb enzyme homologs) and 100 mRNA expressed from the maize ae1 gene were analyzed to establish phylogenetic relationships. Polymorphism of maize ae1 gene different regions was investigated by bioinformatic methods. Modeling of the maize enzyme SBEIIb was performed. Conclusions. According to the results of amino acid sequences of SBEIIb enzyme homologs alignment, it was found that ae1 gene orthologs are present only in monocots, paralogs – in monocots, dicots, and other taxa, including algae and animals. Based on the results of alignment of plants mRNA from which enzyme SBEIIb is translated, maize ae1 gene orthologs and the nearest paralogs encoding starch branching enzymes with chloroplast localization were defined; this suggests a possible origin of ae1 gene due to duplication of the gene encoding the 1,4-alpha-glucan-branching enzyme 2 with chloroplast or amyloplast localization. In the maize ae1 gene structure, regions were found that include polymorphic sites not defined previously. For the polymorphic sites design primers were developed that allowed to differentiate the maize lines. It was determined that the detection of polymorphism in theory can influence the enzyme function and, as a result, change the concentration of amylopectin in maize grain.
显示更多 [+] 显示较少 [-]Rice (<em>Oryza sativa</em> L.) blast resistance genes bioinformatic analysis
2017
Бондаренко, К. В | Сліщук, Г. І | Волкова, Н. Е
Purpose. To investigate rice blast resistance genes polymorphism by using bioinformatic methods. Methods. Global and local nucleotide alignment, phylogenetic analysis, HyPhy test. Results. For Pib gene, numerous single nucleotide substitutions and deletions of 1–3 bp were established. The phylogeny of this gene has been studied and homologues have been found both in various rice species and in other cereals. These sequences can encode proteins that «recognize» the phytopathogens effectors, and can also be associated with resistance to phytopathogens. The Pi4 gene is characterized by single nucleotide substitutions, insertions and deletions; the number of non-synonymous substitutions exceeds the number of synonymous ones. The Pi54 gene variability is significantly lower than that of the Pi4 and Pib genes. The predominant types of polymorphism were single nucleotide substitutions and small-sized indels. It was found that non-synonymous substitutions in Pi54, Pi4 and Pib genes were in close proximity, sometimes forming clusters, while some coding regions were either superconservative or contained predominantly synonymous substitutions. On philodendrograms, cultivated rice samples were clustered with samples of ancestral wild-growing species. Conclusions. Evolution of the rice blast resistance genes Pi4, Pib and Pi54 is characterized by diversification selection. Considering that tense coevolution and significant rate of adaptation and creation of new pathogen races are typical for a plant and a parasite, these genes are subjected to intensive selection aimed at increasing diversity for obtaining the resistance to new races of the pathogen.
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