R. Puzanskiy, Pavel D. Smirnov, Sergey A. Vanisov, Maksim D. Dubrovskiy, A. Shavarda, M. Shishova, V. Yemelyanov
BACKGROUND: The ability of plants to adapt to oxygen deficiency is associated with the presence of various adaptations, many of which are mediated by significant changes of metabolism. These changes allow resistant wetland plants to grow even in oxygen-deficient environment. AIM: The aim of the study was to carry out metabolic profiling of the leaves of the wetland species Epilobium palustre and Epilobium hirsutum, and the mesophyte species Epilobium angustifolium in order to identify the most characteristic metabolome traits of hypoxia-resistant plants. MATERIALS AND METHODS: Metabolite profiling was performed by GC-MS. Statistical analysis of metabolomics data was processed using R 4.2.1 Funny-Looking Kid. RESULTS: The resulting profile included about 360 compounds. 70 of these were identified and 50 compounds were determined to a class. Sugars (64) were the most widely represented in the obtained profiles. 16 amino and 20 carboxylic acids, lipids and secondary compounds have been identified. Significant differences were revealed between the profiles of leaf metabolomes of mesophyte E. angustifolium and hydrophytes E. hirsutum and E. palustre. The mesophyte was characterized by high levels of sugars. The metabolomes of wetland Epilobium species practically did not differ from each other and were characterized by the accumulation of amino acids, including GABA shunt intermediates, dicarboxylic acids of the Krebs cycle, and metabolites of glycolysis and lactic acid fermentation, which reflects the stimulation of anaerobic respiration, nitrogen metabolism, and alternative pathways of NAD(P)H reoxidation in wetland plants. CONCLUSIONS: Traits of metabolic profiles detected in hydrophyte Epilobium species can be used to assess the degree of plant resistance to oxygen deficiency.
{"title":"Metabolite profiling of leaves of three Epilobium species","authors":"R. Puzanskiy, Pavel D. Smirnov, Sergey A. Vanisov, Maksim D. Dubrovskiy, A. Shavarda, M. Shishova, V. Yemelyanov","doi":"10.17816/ecogen114743","DOIUrl":"https://doi.org/10.17816/ecogen114743","url":null,"abstract":"BACKGROUND: The ability of plants to adapt to oxygen deficiency is associated with the presence of various adaptations, many of which are mediated by significant changes of metabolism. These changes allow resistant wetland plants to grow even in oxygen-deficient environment. \u0000AIM: The aim of the study was to carry out metabolic profiling of the leaves of the wetland species Epilobium palustre and Epilobium hirsutum, and the mesophyte species Epilobium angustifolium in order to identify the most characteristic metabolome traits of hypoxia-resistant plants. \u0000MATERIALS AND METHODS: Metabolite profiling was performed by GC-MS. Statistical analysis of metabolomics data was processed using R 4.2.1 Funny-Looking Kid. \u0000RESULTS: The resulting profile included about 360 compounds. 70 of these were identified and 50 compounds were determined to a class. Sugars (64) were the most widely represented in the obtained profiles. 16 amino and 20 carboxylic acids, lipids and secondary compounds have been identified. Significant differences were revealed between the profiles of leaf metabolomes of mesophyte E. angustifolium and hydrophytes E. hirsutum and E. palustre. The mesophyte was characterized by high levels of sugars. The metabolomes of wetland Epilobium species practically did not differ from each other and were characterized by the accumulation of amino acids, including GABA shunt intermediates, dicarboxylic acids of the Krebs cycle, and metabolites of glycolysis and lactic acid fermentation, which reflects the stimulation of anaerobic respiration, nitrogen metabolism, and alternative pathways of NAD(P)H reoxidation in wetland plants. \u0000CONCLUSIONS: Traits of metabolic profiles detected in hydrophyte Epilobium species can be used to assess the degree of plant resistance to oxygen deficiency.","PeriodicalId":11431,"journal":{"name":"Ecological genetics","volume":"83 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2022-12-24","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"74957701","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
AIM: The aim of the study was to carry out species identification of the hybridogenic complex of European water frogs (Pelophylax esculentus complex) in the conditions of transformed biotopes of the south of the Central Russian Upland based on molecular genetic markers MATERIALS AND METHODS: The study included 36 local populations (770 individuals) living in conditions of transformed biotopes of the south of the Central Russian Upland. Identification of cryptic forms was carried out by a Multiplex PCR. Two molecular genetic markers were used for amplification: intron 1 of the SAI-1 DNA serum albumin gene to determine hybrids and cryptic forms, and a fragment of the first subunit of the cytochrome oxidase COI mtDNA gene to determine maternal lines. RESULTS: According to the data obtained, pure R-type population systems predominate (58.33%) in the study region. Mixed RE-type population systems were identified in 14 localities, REL-type in the region is extremely rare and is noted only in one locality. No pure L-type, E-type, or mixed LE-type population systems have been identified. The study revealed a statistically significant (p 0.001) predominance of haplotypes of the Western form (Pelophylax ridibundus). CONCLUSIONS: The data obtained indicate active adaptive changes in the population structure of European water frogs in the study area. Degradation of water bodies caused by abiotic and anthropogenic factors forces amphibians to migrate to neighboring reservoirs in which hybridization of representatives of this complex occurs. Introgressive and hybrid forms of the marsh frog, as well as hybrid edible individuals with greater ecological plasticity and tolerance to anthropogenic pressure, displace populations of Pelophylax lessonae. Based on the above, we consider it necessary to include the pond frog (P. lessonae) in the Red Book of the Belgorod region.
{"title":"Identification of cryptic forms of the hybridogenic complex of European water frogs (Pelophylax esculentus complex) in the conditions of transformed biotopes of the south of the Central Russian Upland based on DNA markers","authors":"Anatoliy S. Barkhatov, E. Snegin","doi":"10.17816/ecogen108544","DOIUrl":"https://doi.org/10.17816/ecogen108544","url":null,"abstract":"AIM: The aim of the study was to carry out species identification of the hybridogenic complex of European water frogs (Pelophylax esculentus complex) in the conditions of transformed biotopes of the south of the Central Russian Upland based on molecular genetic markers \u0000MATERIALS AND METHODS: The study included 36 local populations (770 individuals) living in conditions of transformed biotopes of the south of the Central Russian Upland. Identification of cryptic forms was carried out by a Multiplex PCR. Two molecular genetic markers were used for amplification: intron 1 of the SAI-1 DNA serum albumin gene to determine hybrids and cryptic forms, and a fragment of the first subunit of the cytochrome oxidase COI mtDNA gene to determine maternal lines. \u0000RESULTS: According to the data obtained, pure R-type population systems predominate (58.33%) in the study region. Mixed RE-type population systems were identified in 14 localities, REL-type in the region is extremely rare and is noted only in one locality. No pure L-type, E-type, or mixed LE-type population systems have been identified. The study revealed a statistically significant (p 0.001) predominance of haplotypes of the Western form (Pelophylax ridibundus). \u0000CONCLUSIONS: The data obtained indicate active adaptive changes in the population structure of European water frogs in the study area. Degradation of water bodies caused by abiotic and anthropogenic factors forces amphibians to migrate to neighboring reservoirs in which hybridization of representatives of this complex occurs. Introgressive and hybrid forms of the marsh frog, as well as hybrid edible individuals with greater ecological plasticity and tolerance to anthropogenic pressure, displace populations of Pelophylax lessonae. Based on the above, we consider it necessary to include the pond frog (P. lessonae) in the Red Book of the Belgorod region.","PeriodicalId":11431,"journal":{"name":"Ecological genetics","volume":"22 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2022-12-24","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"82960542","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
D. Isubakova, O. Tsymbal, N. Litviakov, I. Milto, R. M. Takhauov
BACKGROUND: Impaired apoptosis can have serious consequences: the accumulation of mutant cells, the development of teratogenic effects and malignant neoplasms. In this regard, the study of the mechanisms of changes in the activity of apoptosis due to methylation under the influence of long-term irradiation is urgent. AIM: The study of the degree of methylation of gene promoters involved in the induction of apoptosis in the personnel of the Siberian Chemical Plant, exposed to long-term technogenic irradiation of ionizing radiation in the course of their professional activities. MATERIALS AND METHODS: The study was performed on peripheral blood samples of employees of the Siberian Chemical Plant, with a total dose of external exposure from 100 to 300 mSv. Chromosomal aberrations were detected by standard karyotyping of cultured blood lymphocytes. The degree of gene promoters methylation was determined using MethylScreen technology. RESULTS: The degree of gene methylation BIRC2, CASP3, CASP9, CIDEB, CRADD, DAPK1, DFFA, FADD, GADD45A, LTBR, TNFRSF21, TNFRSF25 ranges from 0.31 to 41.75%. A strong negative correlation was found between the degree of methylation of GADD45A (r = 0.7364, р = 0.009) with an increased frequency of aberrant cells, moderate negative correlation GADD45A (r = 0.6347, р = 0.035) with an increased frequency of dicentric chromosomes, moderate negative correlation CASP9 (r = 0.6606, р = 0.026), and strong negative correlation CIDEB (r = 0.7982, р = 0.003) with an increased frequency of chromatid fragments. A moderate negative correlation of the methylation degree of CASP9 (r = 0.6636, р = 0.026), and CIDEB (r = 0.6636, р = 0.026) with the total dose of external exposure was shown. CONCLUSIONS: The decrease in the level of apoptosis at doses of 100300 mSv can be explained by the achievement of the demethylation threshold for the promoters of the proapoptotic genes GADD45A, CASP9, CIDEB. This once again testifies in favor of the threshold model of the dependence of the radiation effect on the radiation dose.
背景:受损的细胞凋亡会产生严重的后果:突变细胞的积累，致畸作用和恶性肿瘤的发展。因此，研究长期照射下甲基化导致细胞凋亡活性变化的机制迫在眉睫。目的:研究西伯利亚化工厂长期电离辐射技术照射人员在职业活动过程中诱导细胞凋亡相关基因启动子的甲基化程度。材料与方法:研究对象为西伯利亚化工厂员工外周血样本，总外照射剂量为100 ~ 300毫西弗。用培养的血淋巴细胞标准核型检测染色体畸变。使用MethylScreen技术测定基因启动子的甲基化程度。结果:BIRC2、CASP3、CASP9、CIDEB、CRADD、DAPK1、DFFA、FADD、GADD45A、LTBR、TNFRSF21、TNFRSF25基因甲基化程度范围为0.31 ~ 41.75%。GADD45A的甲基化程度(r = 0.7364, r = 0.009)与畸变细胞频率的增加呈强负相关，GADD45A的甲基化程度(r = 0.6347, r = 0.035)与双中心染色体频率的增加呈中度负相关，CASP9的甲基化程度(r = 0.6606, r = 0.026)与染色单体片段频率的增加呈中度负相关，CIDEB的甲基化程度(r = 0.7982, r = 0.003)与畸变细胞频率的增加呈强负相关。CASP9甲基化程度(r = 0.6636, r = 0.026)和CIDEB (r = 0.6636, r = 0.026)与总暴露剂量呈中度负相关。结论:100300 mSv剂量下细胞凋亡水平的降低可能与促凋亡基因GADD45A、CASP9、CIDEB启动子的去甲基化阈值达到有关。这再次证明辐射效应与辐射剂量依赖关系的阈值模型是正确的。
{"title":"Relationship between methylation of promoters of apoptosis genes in blood lymphocytes with the frequency of chromosomal aberrations and the dose of radiation","authors":"D. Isubakova, O. Tsymbal, N. Litviakov, I. Milto, R. M. Takhauov","doi":"10.17816/ecogen109119","DOIUrl":"https://doi.org/10.17816/ecogen109119","url":null,"abstract":"BACKGROUND: Impaired apoptosis can have serious consequences: the accumulation of mutant cells, the development of teratogenic effects and malignant neoplasms. In this regard, the study of the mechanisms of changes in the activity of apoptosis due to methylation under the influence of long-term irradiation is urgent. \u0000AIM: The study of the degree of methylation of gene promoters involved in the induction of apoptosis in the personnel of the Siberian Chemical Plant, exposed to long-term technogenic irradiation of ionizing radiation in the course of their professional activities. \u0000MATERIALS AND METHODS: The study was performed on peripheral blood samples of employees of the Siberian Chemical Plant, with a total dose of external exposure from 100 to 300 mSv. Chromosomal aberrations were detected by standard karyotyping of cultured blood lymphocytes. The degree of gene promoters methylation was determined using MethylScreen technology. \u0000RESULTS: The degree of gene methylation BIRC2, CASP3, CASP9, CIDEB, CRADD, DAPK1, DFFA, FADD, GADD45A, LTBR, TNFRSF21, TNFRSF25 ranges from 0.31 to 41.75%. A strong negative correlation was found between the degree of methylation of GADD45A (r = 0.7364, р = 0.009) with an increased frequency of aberrant cells, moderate negative correlation GADD45A (r = 0.6347, р = 0.035) with an increased frequency of dicentric chromosomes, moderate negative correlation CASP9 (r = 0.6606, р = 0.026), and strong negative correlation CIDEB (r = 0.7982, р = 0.003) with an increased frequency of chromatid fragments. A moderate negative correlation of the methylation degree of CASP9 (r = 0.6636, р = 0.026), and CIDEB (r = 0.6636, р = 0.026) with the total dose of external exposure was shown. \u0000CONCLUSIONS: The decrease in the level of apoptosis at doses of 100300 mSv can be explained by the achievement of the demethylation threshold for the promoters of the proapoptotic genes GADD45A, CASP9, CIDEB. This once again testifies in favor of the threshold model of the dependence of the radiation effect on the radiation dose.","PeriodicalId":11431,"journal":{"name":"Ecological genetics","volume":"30 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2022-12-24","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"89161780","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
V. Druzhinin, E. Baranova, P. Demenkov, L. Matskova, S. Paradnikova, V. Volobaev, A. V. Larionov
BACKGROUND: Recent studies show that the bacterial microbiome of the respiratory tract can influence the development of a number of diseases of the human respiratory system. Changes in the composition of the microbiome in patients are associated with dysbiosis, and in addition, many bacteria have a genotoxic potential and can directly or indirectly damage the genome in the cells of the host organism. AIM: The aim of the study was to analyze the composition of the sputum microbiome and its relationship with chromosome damage in the blood leukocytes of patients with chronic dust bronchitis (CDB). MATERIALS AND METHODS: The taxonomic composition of the sputum microbiome of 22 patients with CKD and 22 sputum donors from the control group was studied using next-generation sequencing (NGS) technology of 16S rRNA of bacterial genes. At the same time, the basic frequencies of chromosomal aberrations and micronuclei were determined in blood leukocytes. RESULTS: The sputum microbiome of chronic dust bronchitis patients had a significant reduction in alpha and beta diversity parameters compared to healthy study participants. In addition, an increase in the relative abundance of the genus Streptococcus (29.97 3.03 vs. 18.78 2.47; p = 0.003) was found in the sputum of CP patients compared with the control. Thus, the results of metagenome sequencing indicate a common dysbiotic process with a predominance of one dominant genus of bacteria in this pulmonary pathology. The results of cytogenetic analysis of blood leukocytes showed a significant increase in the proportion of aberrant metaphases in CKD patients compared with healthy donors (3.41% vs. 1.84%; p 0.01) and the absence of significant differences in frequency leukocytes with micronuclei between the compared groups (1.28% vs. 1.11%). Correlation analysis revealed the presence of significant direct relationships between the frequency of aberrant metaphases and the percentage of representatives of the genera Bacteroides in the sputum of patients with chronic dust bronchitis (r = 0.471; p = 0.031); Lachnoanaerobaculum (r = 0.446; p = 0.043) and Alloprevotella (r = 0.444; p = 0.044). Further studies should be devoted to the search for possible mechanisms of influence of these bacteria on clastogenic effects in the cells of the host organism.
背景:最近的研究表明，呼吸道的细菌微生物组可以影响许多人类呼吸系统疾病的发展。患者体内微生物组组成的变化与生态失调有关，此外，许多细菌具有遗传毒性，可以直接或间接损害宿主细胞中的基因组。目的:分析慢性粉尘支气管炎(CDB)患者痰菌群组成及其与血液白细胞染色体损伤的关系。材料与方法:采用细菌基因16S rRNA新一代测序(NGS)技术，研究22例CKD患者和22例对照组痰液微生物组的分类组成。同时测定血液白细胞中染色体畸变和微核的基本频率。结果:与健康研究参与者相比，慢性粉尘支气管炎患者的痰微生物组α和β多样性参数显着减少。此外，链球菌属的相对丰度(29.97 3.03 vs. 18.78 2.47;p = 0.003)。因此，宏基因组测序结果表明，在这种肺部病理中，一种优势菌属的优势是一种常见的生态失调过程。血液白细胞细胞遗传学分析结果显示，与健康供者相比，CKD患者异常中期的比例显著增加(3.41% vs. 1.84%;两组间微核白细胞频率差异无统计学意义(1.28% vs. 1.11%)。相关性分析显示，慢性粉尘支气管炎患者痰液中拟杆菌属代表菌的比例与异常中期的发生频率有显著的直接关系(r = 0.471;P = 0.031);厌氧菌群(r = 0.446;p = 0.043)和异丙普氏菌(r = 0.444;P = 0.044)。进一步的研究应致力于寻找这些细菌对宿主细胞中致裂作用的可能影响机制。
{"title":"Chronic dust bronchitis: composition of the sputum bacterial microbiome and its association with chromosome damage in blood lymphocytes","authors":"V. Druzhinin, E. Baranova, P. Demenkov, L. Matskova, S. Paradnikova, V. Volobaev, A. V. Larionov","doi":"10.17816/ecogen108807","DOIUrl":"https://doi.org/10.17816/ecogen108807","url":null,"abstract":"BACKGROUND: Recent studies show that the bacterial microbiome of the respiratory tract can influence the development of a number of diseases of the human respiratory system. Changes in the composition of the microbiome in patients are associated with dysbiosis, and in addition, many bacteria have a genotoxic potential and can directly or indirectly damage the genome in the cells of the host organism. \u0000AIM: The aim of the study was to analyze the composition of the sputum microbiome and its relationship with chromosome damage in the blood leukocytes of patients with chronic dust bronchitis (CDB). \u0000MATERIALS AND METHODS: The taxonomic composition of the sputum microbiome of 22 patients with CKD and 22 sputum donors from the control group was studied using next-generation sequencing (NGS) technology of 16S rRNA of bacterial genes. At the same time, the basic frequencies of chromosomal aberrations and micronuclei were determined in blood leukocytes. \u0000RESULTS: The sputum microbiome of chronic dust bronchitis patients had a significant reduction in alpha and beta diversity parameters compared to healthy study participants. In addition, an increase in the relative abundance of the genus Streptococcus (29.97 3.03 vs. 18.78 2.47; p = 0.003) was found in the sputum of CP patients compared with the control. Thus, the results of metagenome sequencing indicate a common dysbiotic process with a predominance of one dominant genus of bacteria in this pulmonary pathology. The results of cytogenetic analysis of blood leukocytes showed a significant increase in the proportion of aberrant metaphases in CKD patients compared with healthy donors (3.41% vs. 1.84%; p 0.01) and the absence of significant differences in frequency leukocytes with micronuclei between the compared groups (1.28% vs. 1.11%). Correlation analysis revealed the presence of significant direct relationships between the frequency of aberrant metaphases and the percentage of representatives of the genera Bacteroides in the sputum of patients with chronic dust bronchitis (r = 0.471; p = 0.031); Lachnoanaerobaculum (r = 0.446; p = 0.043) and Alloprevotella (r = 0.444; p = 0.044). Further studies should be devoted to the search for possible mechanisms of influence of these bacteria on clastogenic effects in the cells of the host organism.","PeriodicalId":11431,"journal":{"name":"Ecological genetics","volume":"890 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2022-12-24","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"77013175","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
BACKGROUND: The infrared reflection spectroscopy application method for rapid assessment of biochemical parameters in various types of oats is shown. On the basis of the biochemical data obtained in the laboratory of VIR, calibration models of protein, oil and starch content were constructed. AIM: The aim of the study is to develop an express method of near-infrared spectroscopy (NIRS) spectroscopy to determine the main biochemical parameters in oat seeds and to build calibration models for the MATRIX-I IR analyser to quantify the mass fraction of protein, oil and starch in oat seeds based on data obtained by traditional methods. MATERIALS AND METHODS: Biochemical quality indicators (protein, oil, starch) were studied on seeds of filmy oats (Avena sativa L.) grown in 20152016 in the North-Western Region of the Russian Federation. Calibration models for the determination of protein, oil and starch in oat seeds (98 samples, harvest 20142015) were developed for the MATRIX-I IR analyzer by Bruker Optics (Germany). Values obtained by traditional chemical methods of analysis were used to construct calibration models. Oat seed oil was determined by the Soxlet method, protein by the Kjeldahl method, starch by the Evers polarimetric method. All indicators were recalculated for dry weight. RESULTS AND CONCLUSION: The reliability of the developed models was checked by the results of protein, oil and starch determination in the seeds of the test batch according to the indicator of the calibration correctness. The data obtained using the calibration curve on the MATRIX-I device had no significant differences with the results of chemical studies. Therefore, calibration can be used for screening analysis for protein, oil and starch content in oat samples. This method allows you to save valuable material, increase labour productivity due to the speed of obtaining data, does not require reagents and is safe.
{"title":"Application of the IR spectrometry method in the screening study of various oat species","authors":"V. Khoreva, V. S. Popov, N. Kon’kova","doi":"10.17816/ecogen108503","DOIUrl":"https://doi.org/10.17816/ecogen108503","url":null,"abstract":"BACKGROUND: The infrared reflection spectroscopy application method for rapid assessment of biochemical parameters in various types of oats is shown. On the basis of the biochemical data obtained in the laboratory of VIR, calibration models of protein, oil and starch content were constructed. \u0000AIM: The aim of the study is to develop an express method of near-infrared spectroscopy (NIRS) spectroscopy to determine the main biochemical parameters in oat seeds and to build calibration models for the MATRIX-I IR analyser to quantify the mass fraction of protein, oil and starch in oat seeds based on data obtained by traditional methods. \u0000MATERIALS AND METHODS: Biochemical quality indicators (protein, oil, starch) were studied on seeds of filmy oats (Avena sativa L.) grown in 20152016 in the North-Western Region of the Russian Federation. Calibration models for the determination of protein, oil and starch in oat seeds (98 samples, harvest 20142015) were developed for the MATRIX-I IR analyzer by Bruker Optics (Germany). Values obtained by traditional chemical methods of analysis were used to construct calibration models. Oat seed oil was determined by the Soxlet method, protein by the Kjeldahl method, starch by the Evers polarimetric method. All indicators were recalculated for dry weight. \u0000RESULTS AND CONCLUSION: The reliability of the developed models was checked by the results of protein, oil and starch determination in the seeds of the test batch according to the indicator of the calibration correctness. The data obtained using the calibration curve on the MATRIX-I device had no significant differences with the results of chemical studies. Therefore, calibration can be used for screening analysis for protein, oil and starch content in oat samples. This method allows you to save valuable material, increase labour productivity due to the speed of obtaining data, does not require reagents and is safe.","PeriodicalId":11431,"journal":{"name":"Ecological genetics","volume":"96 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2022-12-24","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"85781870","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
E. V. Markova, A. V. Chirinskaite, J. Sopova, E. Leonova
In the modern world, due to the overconsumption of sugar-containing products, the problem of obesity is relevant. Among the many sweeteners that minimize sugar intake, a group of sweet-tasting proteins is up-and-coming. Brazzein is the smallest of the sweet proteins (54 aa, 6473 Da), and it is also safe for obese and diabetic people since it does not affect blood sugar and insulin levels. Brazzein has high thermal stability over a wide pH range: from 2 to 8 [1]. To increase the level of sweetness of brazzein, mutant variants of this protein were created through site-directed mutagenesis, the sweetest of which is triple mutant H31R/E36D/E41A, which is 22,500 times sweeter than sucrose [2]. Since the content of brazzein in the fruits of the natural source (Pentadiplandra brazzeana) is extremely low (0.2%), various methods have been developed to obtain brazzein using heterologous expression systems, which used as producers: bacteria (Escherichia coli, Lactococcus lactis), yeast (Pichia pastoris, Kluyveromyces lactis, Saccharomyces cerevisiae), plants (Zea mays, Oryza sativa, Lactuca sativa, Nicotiana tabacum) and animals (Mus musculus) [35]. Despite the short peptide sequence, the industrial production of recombinant protein faced several problems, including low protein yield (e.g in mouse milk it was detectable on western blot analysis only) and loss of sweetness. Аn extremely relevant and promising way to obtain recombinant brazzein is the optimization of extracellular expression in bakers yeasts with the GRAS (Generally recognized as safe) status, since the safety of these microorganisms for human health can potentially significantly reduce the number of brazzein purification steps and thereby reduce its cost to consumers.
{"title":"The sweet protein brazzein as a promising natural sweetener","authors":"E. V. Markova, A. V. Chirinskaite, J. Sopova, E. Leonova","doi":"10.17816/ecogen112373","DOIUrl":"https://doi.org/10.17816/ecogen112373","url":null,"abstract":"In the modern world, due to the overconsumption of sugar-containing products, the problem of obesity is relevant. Among the many sweeteners that minimize sugar intake, a group of sweet-tasting proteins is up-and-coming. Brazzein is the smallest of the sweet proteins (54 aa, 6473 Da), and it is also safe for obese and diabetic people since it does not affect blood sugar and insulin levels. Brazzein has high thermal stability over a wide pH range: from 2 to 8 [1]. To increase the level of sweetness of brazzein, mutant variants of this protein were created through site-directed mutagenesis, the sweetest of which is triple mutant H31R/E36D/E41A, which is 22,500 times sweeter than sucrose [2]. Since the content of brazzein in the fruits of the natural source (Pentadiplandra brazzeana) is extremely low (0.2%), various methods have been developed to obtain brazzein using heterologous expression systems, which used as producers: bacteria (Escherichia coli, Lactococcus lactis), yeast (Pichia pastoris, Kluyveromyces lactis, Saccharomyces cerevisiae), plants (Zea mays, Oryza sativa, Lactuca sativa, Nicotiana tabacum) and animals (Mus musculus) [35]. Despite the short peptide sequence, the industrial production of recombinant protein faced several problems, including low protein yield (e.g in mouse milk it was detectable on western blot analysis only) and loss of sweetness. Аn extremely relevant and promising way to obtain recombinant brazzein is the optimization of extracellular expression in bakers yeasts with the GRAS (Generally recognized as safe) status, since the safety of these microorganisms for human health can potentially significantly reduce the number of brazzein purification steps and thereby reduce its cost to consumers.","PeriodicalId":11431,"journal":{"name":"Ecological genetics","volume":"39 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2022-12-08","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"79367798","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Anton I. Degtyarenko, Varvara D Stepochkina, Y. Shkryl
Plant genetic modification in order to increase their tolerance to various abiotic stresses has been of exceptional importance in recent years. Heterologous expression of glycine betaine (GB) biosynthetic genes leads to increased salt and drought tolerance in various plant species by maintaining the osmotic balance with the environment and stabilizing the quaternary structure of complex proteins. However, GB biosynthesis in transgenic plants is limited by choline availability. Members of the Plumbaginaceae family accumulate -alanine betaine (AB) instead [1]. The synthesis of AB is not limited by the availability of choline, as it follows the methylation pathway of the aproteinogenic amino acid -alanine. For the first time, we have generated Nicotiana tabacum plants expressing the -alanine N-methyltransferase (LlBANMT) gene of Limonium latifolium. Transgenic plants were much less affected by such abiotic stresses as increased salinity, excessive illumination, and low temperature. The experimental Nicotiana tabacum lines had lower rates of chlorophyll degradation under stress conditions compared to the control plants. LlBANMT expression also resulted in less biomass loss under stress conditions, which was associated with higher activities of reactive oxygen species detoxification systems and healthier cell membranes. The presented data demonstrate for the first time the protective properties of LlBANMT heterologous expression and shed light on the mechanisms of its action.
{"title":"Heterologous expression of β-alanine betaine biosynthesis gene increases Nicotiana tabacum resistance to abiotic stresses","authors":"Anton I. Degtyarenko, Varvara D Stepochkina, Y. Shkryl","doi":"10.17816/ecogen112339","DOIUrl":"https://doi.org/10.17816/ecogen112339","url":null,"abstract":"Plant genetic modification in order to increase their tolerance to various abiotic stresses has been of exceptional importance in recent years. Heterologous expression of glycine betaine (GB) biosynthetic genes leads to increased salt and drought tolerance in various plant species by maintaining the osmotic balance with the environment and stabilizing the quaternary structure of complex proteins. However, GB biosynthesis in transgenic plants is limited by choline availability. Members of the Plumbaginaceae family accumulate -alanine betaine (AB) instead [1]. The synthesis of AB is not limited by the availability of choline, as it follows the methylation pathway of the aproteinogenic amino acid -alanine. \u0000For the first time, we have generated Nicotiana tabacum plants expressing the -alanine N-methyltransferase (LlBANMT) gene of Limonium latifolium. Transgenic plants were much less affected by such abiotic stresses as increased salinity, excessive illumination, and low temperature. The experimental Nicotiana tabacum lines had lower rates of chlorophyll degradation under stress conditions compared to the control plants. LlBANMT expression also resulted in less biomass loss under stress conditions, which was associated with higher activities of reactive oxygen species detoxification systems and healthier cell membranes. The presented data demonstrate for the first time the protective properties of LlBANMT heterologous expression and shed light on the mechanisms of its action.","PeriodicalId":11431,"journal":{"name":"Ecological genetics","volume":"9 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2022-12-08","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"90806077","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Radish (Raphanus sativus L.) is an agronomically important root crop belonging to the Brassicaceae family. A genetic collection of inbred radish lines, which was based on contrasting genetically determined traits including trait ability to develop tumors, was created at St. Petersburg State University in the middle of the 20th century [1]. Full genetics network controlling this trait still remains unclear [2], and elucidation of its mechanisms can help reveal the key regulators of systemic mechanisms that control cell proliferation and differentiation. The purpose of this work is to identify single nucleotide polymorphisms (SNPs) and InDels in genes which are candidates for participation in the tumor development process within in the tumor-forming radish line compared to the non-tumor radish line. We have assembled the genomes of two lines contrasting in the ability to tumorigenesis, annotated them, aligned sequences per assembly, identified candidate genes and differences in the structure of these genes in contrasting radish lines using bioinformatics tools. Bioinformatics data were confirmed by the sequencing by Sanger method. As a result, in the tumor-forming radish line we identified 151 genes with InDels in their coding regions, which led to various variants of frameshift. Moreover, we detected 39 genes with single nucleotide substitutions (SNPs). According to the gene pathway enrichment analysis, the corresponding genes were classified into the several groups. Thus, the data obtained will allow us to clarify in more detail the genetic control mechanisms of the tumor development in radish. The work was supported by the Ministry of Science and Higher Education of the Russian Federation in accordance with the agreement No. 075-15-2022-322.
{"title":"Identification of SNPs and InDels probably associated with the development of spontaneous tumors in radish (Raphanus sativus L.)","authors":"Ksenia A. Kuznetsova, I. Dodueva, L. Lutova","doi":"10.17816/ecogen112307","DOIUrl":"https://doi.org/10.17816/ecogen112307","url":null,"abstract":"Radish (Raphanus sativus L.) is an agronomically important root crop belonging to the Brassicaceae family. A genetic collection of inbred radish lines, which was based on contrasting genetically determined traits including trait ability to develop tumors, was created at St. Petersburg State University in the middle of the 20th century [1]. Full genetics network controlling this trait still remains unclear [2], and elucidation of its mechanisms can help reveal the key regulators of systemic mechanisms that control cell proliferation and differentiation. \u0000The purpose of this work is to identify single nucleotide polymorphisms (SNPs) and InDels in genes which are candidates for participation in the tumor development process within in the tumor-forming radish line compared to the non-tumor radish line. \u0000We have assembled the genomes of two lines contrasting in the ability to tumorigenesis, annotated them, aligned sequences per assembly, identified candidate genes and differences in the structure of these genes in contrasting radish lines using bioinformatics tools. Bioinformatics data were confirmed by the sequencing by Sanger method. \u0000As a result, in the tumor-forming radish line we identified 151 genes with InDels in their coding regions, which led to various variants of frameshift. Moreover, we detected 39 genes with single nucleotide substitutions (SNPs). According to the gene pathway enrichment analysis, the corresponding genes were classified into the several groups. \u0000Thus, the data obtained will allow us to clarify in more detail the genetic control mechanisms of the tumor development in radish. \u0000The work was supported by the Ministry of Science and Higher Education of the Russian Federation in accordance with the agreement No. 075-15-2022-322.","PeriodicalId":11431,"journal":{"name":"Ecological genetics","volume":"351 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2022-12-08","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"76579155","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
E. Dolgikh, Elizaveta S. Kantsurova, Alina M. Dymo
Plants interact with a wide range of soil microorganisms, while interactions with symbiotic microorganisms are the most important for them. Symbiosis with nitrogen-fixing nodule bacteria provides a significant advantage in the existence of plants in nitrogen-poor soils. Since only plants from the Fabales and Rosales (a single representative Parasponia) enter into symbiosis with nodule bacteria, an idea about expanding the number of plants entering into such interactions became popular. To solve the problem of constructing new symbiotic systems, it is necessary to provide recognition of the symbiont (by transferring receptor genes into non-legume plants), morphogenesis of the nodule, and its infection. We have managed to introduce the genes encoding receptors to surface components of rhizobia into the non-legume plants, and it provided increased colonization by nodule bacteria. It may improve the growth and development of non-legume plants and ensure their greater resistance to phytopathogens. Genome analysis of non-legume plant hop Humulus lupulus showed the existence of genes involved in symbiosis regulation, but some important regulators such as gene encoding NIN transcription factor were lost. Using genetic engineering approaches, the hop plants carrying the gene encoding NIN transcription factor were created. Analysis of these plants may provide important information about regulation of organogenesis in non-legume plants. The work was financially supported by RSF 21-16-00106.
{"title":"The development of approaches to create new symbiotic systems","authors":"E. Dolgikh, Elizaveta S. Kantsurova, Alina M. Dymo","doi":"10.17816/ecogen112392","DOIUrl":"https://doi.org/10.17816/ecogen112392","url":null,"abstract":"Plants interact with a wide range of soil microorganisms, while interactions with symbiotic microorganisms are the most important for them. Symbiosis with nitrogen-fixing nodule bacteria provides a significant advantage in the existence of plants in nitrogen-poor soils. Since only plants from the Fabales and Rosales (a single representative Parasponia) enter into symbiosis with nodule bacteria, an idea about expanding the number of plants entering into such interactions became popular. To solve the problem of constructing new symbiotic systems, it is necessary to provide recognition of the symbiont (by transferring receptor genes into non-legume plants), morphogenesis of the nodule, and its infection. \u0000We have managed to introduce the genes encoding receptors to surface components of rhizobia into the non-legume plants, and it provided increased colonization by nodule bacteria. It may improve the growth and development of non-legume plants and ensure their greater resistance to phytopathogens. Genome analysis of non-legume plant hop Humulus lupulus showed the existence of genes involved in symbiosis regulation, but some important regulators such as gene encoding NIN transcription factor were lost. Using genetic engineering approaches, the hop plants carrying the gene encoding NIN transcription factor were created. Analysis of these plants may provide important information about regulation of organogenesis in non-legume plants. \u0000The work was financially supported by RSF 21-16-00106.","PeriodicalId":11431,"journal":{"name":"Ecological genetics","volume":"44 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2022-12-08","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"76205611","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
In this work we knocked out the LTS3 gene of the microalgae Chlamydomonas reinhardtii using the TIM technique optimized for the available equipment. We achieved transformation efficiency of 68.8%, knockout of this gene lead to the death of C. reinhardtii cells after several division cycles. The creation and study of genetically modified organisms in fundamental research allows a deeper understanding of the basic processes in the cells with the prospect of further applying this knowledge in practice. Microalgae are an interesting object for genetic engineering because of the great prospects for their application in biotechnology, but in almost every case it is necessary to develop new strategies and transformation methods for the introduction of genetic constructs into the cell. CRISPR/Cas revolutionized the field of genome editing due to its simplicity, efficiency and accuracy compared to previously used methods, which over time simplified the development of protocols [1]. Currently, the most effective method of transformation is TIM (Targeted Insertional Mutagenesis) [2], developed for the microalgae Chlamydomonas reinhardtii P.A. Dang. model object of photosynthesis genetics. To test and optimize the TIM technique [2] in our lab, we carried out a knockout of the LTS3 gene, a transcriptional activator of chlorophyll biosynthesis genes in heterotrophic conditions [3]. We used glass beads agitation and electroporation (Gene Pulser Xcell, Bio-Rad, USA) methods in order to introduce into C. reinhardtii cells of the CC-125 (wt, mt+) strain the ribonucleoprotein complex SpCas9/sgRNA and double-stranded donor DNA with paromomycin resistance gene. The effectiveness of transformation varied from 10.6% to 68.8%. Probably, the LTS3 gene product plays a key role in the pathway of chlorophyll biosynthesis, since its knockout led to the death of C. reinhardtii cells after several division cycles. The transformation protocol optimized for the equipment available in our lab can be further refined and used to study the functions of other C. reinhardtii genes.
{"title":"Optimization of CRISPR/Cas9 method for transgenesis of model microalgae Chlamydomonas reinhardtii","authors":"Pavel A. Virolainen, E. Chekunova","doi":"10.17816/ecogen112332","DOIUrl":"https://doi.org/10.17816/ecogen112332","url":null,"abstract":"In this work we knocked out the LTS3 gene of the microalgae Chlamydomonas reinhardtii using the TIM technique optimized for the available equipment. We achieved transformation efficiency of 68.8%, knockout of this gene lead to the death of C. reinhardtii cells after several division cycles. \u0000The creation and study of genetically modified organisms in fundamental research allows a deeper understanding of the basic processes in the cells with the prospect of further applying this knowledge in practice. Microalgae are an interesting object for genetic engineering because of the great prospects for their application in biotechnology, but in almost every case it is necessary to develop new strategies and transformation methods for the introduction of genetic constructs into the cell. CRISPR/Cas revolutionized the field of genome editing due to its simplicity, efficiency and accuracy compared to previously used methods, which over time simplified the development of protocols [1]. Currently, the most effective method of transformation is TIM (Targeted Insertional Mutagenesis) [2], developed for the microalgae Chlamydomonas reinhardtii P.A. Dang. model object of photosynthesis genetics. \u0000To test and optimize the TIM technique [2] in our lab, we carried out a knockout of the LTS3 gene, a transcriptional activator of chlorophyll biosynthesis genes in heterotrophic conditions [3]. \u0000We used glass beads agitation and electroporation (Gene Pulser Xcell, Bio-Rad, USA) methods in order to introduce into C. reinhardtii cells of the CC-125 (wt, mt+) strain the ribonucleoprotein complex SpCas9/sgRNA and double-stranded donor DNA with paromomycin resistance gene. \u0000The effectiveness of transformation varied from 10.6% to 68.8%. Probably, the LTS3 gene product plays a key role in the pathway of chlorophyll biosynthesis, since its knockout led to the death of C. reinhardtii cells after several division cycles. \u0000The transformation protocol optimized for the equipment available in our lab can be further refined and used to study the functions of other C. reinhardtii genes.","PeriodicalId":11431,"journal":{"name":"Ecological genetics","volume":"83 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2022-12-08","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"74548406","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
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