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Nicotine exhibits obvious heterosis, which can be used to create Nicotiana tabacum L. (tobacco) varieties with varying nicotine content. However, the reasons for the formation of nicotine heterosis and its relationship to nicotine transport and accumulation remain unknown. This study conducted a comprehensive analysis of six tobacco hybrids with varying heterosis levels and their parent materials from various aspects, such as phenotype, physiology, and transcriptomics. The results showed that the direct path coefficient of transport heterosis to nicotine heterosis was highest in hybrids, at 0.98, and a highly significant positive correlation between the two. The plant height, thick stalk circumference, large flow of tissue fluid in the stalk, and high nicotine concentration of tobacco were the underlying factors that led to the strong nicotine transport capacity of hybrids. The formation of nicotine transport heterosis in hybrids was mainly influenced by non-additive gene effects (accounting for 89.93%), with over-dominant effects playing a dominant role (accounting for 58.79%). Among non-additive expression DEGs, nicotine transporter related multi antimicrobial extrusion protein, drug/metabolite transporter, ABC family transporter, and glutathione S-transferase were significantly upregulated in hybrid strains. The RT-qPCR results indicated that these genes related nicotine transport also exhibited heterosis at the expression level. Our results revealed that the formation of nicotine heterosis is mainly achieved by enhancing the nicotine transport capacity in hybrids. The results are not only beneficial for promoting the theoretical study of nicotine heterosis in tobacco and the breeding and utilization of hybrids, but are also of great significance for guiding nicotine production and promoting its multipurpose utilization. [ABSTRACT FROM AUTHOR]

Heterosis, recognized for improving crop performance, especially in the first filial (F1) generation, remains an area of significant study in the tobacco industry. The low utilization of leaf veins in tobacco contributes to economic inefficiency and resource waste. Despite the positive impacts of heterosis on crop genetics, investigations into leaf-vein ratio heterosis in tobacco have been lacking. Understanding the mechanisms underlying negative heterosis in leaf vein ratio at the molecular level is crucial for advancing low vein ratio leaf breeding research. This study involved 12 hybrid combinations and their parental lines to explore heterosis associated with leaf vein ratios. The hybrids displayed diverse patterns of positive or negative leaf vein ratio heterosis across different developmental stages. Notably, the F1 hybrid (G70 × Qinggeng) consistently exhibited substantial negative heterosis, reaching a maximum of -19.79% 80 days after transplanting. A comparative transcriptome analysis revealed that a significant proportion of differentially expressed genes (DEGs), approximately 39.04% and 23.73%, exhibited dominant and over-dominant expression patterns, respectively. These findings highlight the critical role of non-additive gene expression, particularly the dominance pattern, in governing leaf vein ratio heterosis. The non-additive genes, largely associated with various GO terms such as response to abiotic stimuli, galactose metabolic process, plant-type cell wall organization, auxin-activated signaling pathway, hydrolase activity, and UDP-glycosyltransferase activity, were identified. Furthermore, KEGG enrichment analysis unveiled their involvement in phenylpropanoid biosynthesis, galactose metabolism, plant hormone signal transduction, glutathione metabolism, MAPK signaling pathway, starch, and sucrose metabolism. Among the non-additive genes, we identified some genes related to leaf development, leaf size, leaf senescence, and cell wall extensibility that showed significantly lower expression in F1 than in its parents. These results indicate that the non-additive expression of genes plays a key role in the heterosis of the leaf vein ratio in tobacco. This study marks the first exploration into the molecular mechanisms governing leaf vein ratio heterosis at the transcriptome level. These findings significantly contribute to understanding leaf vein ratios in tobacco breeding strategies. [ABSTRACT FROM AUTHOR]

Hybrid cultivars could be beneficial to achieve high leaf yields while maintaining good quality properties in oriental tobacco. Identification of parents constitutes a major part of a hybrid breeding program. The aim of the present study was to determine the relationship between the genetic distance of parents determined by simple sequence repeats (SSR) markers and heterosis levels in hybrids produced from those parents for leaf yield and quality properties in oriental tobacco. Twenty-one hybrids produced by half diallel crossing among seven tobacco genotypes used in oriental tobacco production in Turkey were grown along with their parents in three locations in 2012 and 2013. Twenty-nine SSR markers were used to determine genetic distances among seven tobacco genotypes. A total of 80 alleles were produced by all twenty-nine markers. Average number of observed alleles per polymorphic marker was 2.96. Twenty-seven of 80 alleles were observed in only one of the seven parents. The polymorphic information content of markers varied from 0.215 to 0.810 (average 0.480). Mid-parent heterosis levels ranged from 18.03 to 42.00% for leaf number, between -19.75 and 38.06% for leaf width, between -17.51 and 36.25% for leaf length, between -34.38 and 76.12% for leaf yield, between -78.30 and 154.01% for sugar content and between -45.40 and 143.29% for nicotine content. Heterosis levels were correlated with genetic distances between parents for leaf number in Erbaa 2012 and Tokat 2012 locations, for leaf width in Erbaa 2013 location, for leaf length in Erbaa 2013 location, and for leaf yield in Tokat 2012 location only. SSR markers were very effective to determine genetic distance of oriental tobacco, and only two markers could distinguish all seven genotypes used in the study. The findings indicated that genetic distance determined by SSR markers used in the present study is not sufficient to predict hybrid performance in oriental tobacco. [ABSTRACT FROM AUTHOR]

Abstract Nicotine exhibits obvious heterosis, which can be used to create Nicotiana tabacum L. (tobacco) varieties with varying nicotine content. However, the reasons for the formation of nicotine heterosis and its relationship to nicotine transport and accumulation remain unknown. This study conducted a comprehensive analysis of six tobacco hybrids with varying heterosis levels and their parent materials from various aspects, such as phenotype, physiology, and transcriptomics. The results showed that the direct path coefficient of transport heterosis to nicotine heterosis was highest in hybrids, at 0.98, and a highly significant positive correlation between the two. The plant height, thick stalk circumference, large flow of tissue fluid in the stalk, and high nicotine concentration of tobacco were the underlying factors that led to the strong nicotine transport capacity of hybrids. The formation of nicotine transport heterosis in hybrids was mainly influenced by non-additive gene effects (accounting for 89.93%), with over-dominant effects playing a dominant role (accounting for 58.79%). Among non-additive expression DEGs, nicotine transporter related multi antimicrobial extrusion protein, drug/metabolite transporter, ABC family transporter, and glutathione S-transferase were significantly upregulated in hybrid strains. The RT-qPCR results indicated that these genes related nicotine transport also exhibited heterosis at the expression level. Our results revealed that the formation of nicotine heterosis is mainly achieved by enhancing the nicotine transport capacity in hybrids. The results are not only beneficial for promoting the theoretical study of nicotine heterosis in tobacco and the breeding and utilization of hybrids, but are also of great significance for guiding nicotine production and promoting its multipurpose utilization.

Abstract Heterosis, recognized for improving crop performance, especially in the first filial (F1) generation, remains an area of significant study in the tobacco industry. The low utilization of leaf veins in tobacco contributes to economic inefficiency and resource waste. Despite the positive impacts of heterosis on crop genetics, investigations into leaf-vein ratio heterosis in tobacco have been lacking. Understanding the mechanisms underlying negative heterosis in leaf vein ratio at the molecular level is crucial for advancing low vein ratio leaf breeding research. This study involved 12 hybrid combinations and their parental lines to explore heterosis associated with leaf vein ratios. The hybrids displayed diverse patterns of positive or negative leaf vein ratio heterosis across different developmental stages. Notably, the F1 hybrid (G70 × Qinggeng) consistently exhibited substantial negative heterosis, reaching a maximum of -19.79% 80 days after transplanting. A comparative transcriptome analysis revealed that a significant proportion of differentially expressed genes (DEGs), approximately 39.04% and 23.73%, exhibited dominant and over-dominant expression patterns, respectively. These findings highlight the critical role of non-additive gene expression, particularly the dominance pattern, in governing leaf vein ratio heterosis. The non-additive genes, largely associated with various GO terms such as response to abiotic stimuli, galactose metabolic process, plant-type cell wall organization, auxin-activated signaling pathway, hydrolase activity, and UDP-glycosyltransferase activity, were identified. Furthermore, KEGG enrichment analysis unveiled their involvement in phenylpropanoid biosynthesis, galactose metabolism, plant hormone signal transduction, glutathione metabolism, MAPK signaling pathway, starch, and sucrose metabolism. Among the non-additive genes, we identified some genes related to leaf development, leaf size, leaf senescence, and cell wall extensibility that showed significantly lower expression in F1 than in its parents. These results indicate that the non-additive expression of genes plays a key role in the heterosis of the leaf vein ratio in tobacco. This study marks the first exploration into the molecular mechanisms governing leaf vein ratio heterosis at the transcriptome level. These findings significantly contribute to understanding leaf vein ratios in tobacco breeding strategies.

Tobacco is an important non-food crop with many applications, which is widely cultivated worldwide. Genetic linkage maps play an important role in QTL mapping, gene cloning, marker assistant selection, and genome assembly However, due to the narrow genetic background and large genome of tobacco, the research on the tobacco genetic map lags behind. At present, although several linkage maps have been constructed based on SSR (simple sequence repeat) markers, different SSR markers limit their widespread applications. In order to construct an integrated linkage map, we identified the redundancy of tobacco SSR markers, and constructed a high-quality genetic map by using the integration method based on four tobacco genetic maps. Finally, 20 redundant markers were found from 3.354 SSR markers, and the integrated high-density genetic linkage map (IHD) had a total of 3.377 loci with a length of 2.489.82 cM and a density of 1.36 locus/cM. The high-density integrated genetic map will have great application in the field of QTL mapping and marker-assisted breeding in tobacco. [ABSTRACT FROM AUTHOR]

Abstract Background The dirigent (DIR) genes encode proteins that act as crucial regulators of plant lignin biosynthesis. In Solanaceae species, members of the DIR gene family are intricately related to plant growth and development, playing a key role in responding to various biotic and abiotic stresses. It will be of great application significance to analyze the DIR gene family and expression profile under various pathogen stresses in Solanaceae species. Results A total of 57 tobacco NtDIRs and 33 potato StDIRs were identified based on their respective genome sequences. Phylogenetic analysis of DIR genes in tobacco, potato, eggplant and Arabidopsis thaliana revealed three distinct subgroups (DIR-a, DIR-b/d and DIR-e). Gene structure and conserved motif analysis showed that a high degree of conservation in both exon/intron organization and protein motifs among tobacco and potato DIR genes, especially within members of the same subfamily. Total 8 pairs of tandem duplication genes (3 pairs in tobacco, 5 pairs in potato) and 13 pairs of segmental duplication genes (6 pairs in tobacco, 7 pairs in potato) were identified based on the analysis of gene duplication events. Cis-regulatory elements of the DIR promoters participated in hormone response, stress responses, circadian control, endosperm expression, and meristem expression. Transcriptomic data analysis under biotic stress revealed diverse response patterns among DIR gene family members to pathogens, indicating their functional divergence. After 96 h post-inoculation with Ralstonia solanacearum L. (Ras), tobacco seedlings exhibited typical symptoms of tobacco bacterial wilt. The qRT-PCR analysis of 11 selected NtDIR genes displayed differential expression pattern in response to the bacterial pathogen Ras infection. Using line 392278 of potato as material, typical symptoms of potato late blight manifested on the seedling leaves under Phytophthora infestans infection. The qRT-PCR analysis of 5 selected StDIR genes showed up-regulation in response to pathogen infection. Notably, three clustered genes (NtDIR2, NtDIR4, StDIR3) exhibited a robust response to pathogen infection, highlighting their essential roles in disease resistance. Conclusion The genome-wide identification, evolutionary analysis, and expression profiling of DIR genes in response to various pathogen infection in tobacco and potato have provided valuable insights into the roles of these genes under various stress conditions. Our results could provide a basis for further functional analysis of the DIR gene family under pathogen infection conditions.

Mulching is one of the most recommended practices in agriculture due to its positive effects on the plant/soil system. Very few experiments have been conducted to date to investigate the effect of mulching, with both organic and inorganic materials, on tobacco. The main aim of this study was to test the synergic effect of a soil-biodegradable (according to standard EN17033) mulching film (the commercial Mater-Bi®, Novamont SpA, Novara, Italy) and drip irrigation (M-D) compared with that of bare soil and sprinkler/drip irrigation (first/second part of the growing season; BS-SD) on a tobacco crop (Nicotiana tabacum L., flue-cured Virginia) grown in the Tiber Valley (the tobacco cultivation district of Central Italy). BS-SD represents the standard practice applied by tobacco growers in the study area. The plants grown under the M-D management system grew more and developed faster than the plants grown under BS-SD conditions. Under the M-D system, yields increased in comparison with the BS-SD conditions (+29%, on average). The gross revenue obtained via the M-D-cured products also increased (+63%, on average) thanks to higher prices assigned by expert evaluators on the basis of the extrinsic quality traits (color, structure and texture, degree of ripeness, elasticity, lamina integrity, handling defects, and vein incidence). The economic value of the cured products increased with the leaf crowns; it was the lowest in the basal (B) leaves and the highest in the middle-upper (MU) leaves. The intrinsic quality traits of the cured leaves (total N and nitrate contents, alkaloids, and reducing sugars) also confirmed that the best quality was found in the M-D-cured products, as determined by expert evaluation. Interestingly, the reducing sugar (RS) contents of tobacco obtained using the M-D management system were 2.5-, 1.1-, and 0.9-fold greater than those under the BS-SD conditions (B, M, and MU products, respectively). An additional commercial value of the cured products was thus obtained with the M-D crop management system due to RS, an intrinsic quality trait considered by manufacturing industries. [ABSTRACT FROM AUTHOR]

The E3 enzyme in the UPS pathway is a crucial factor for inhibiting substrate specificity. In Solanaceae, the U-box E3 ubiquitin ligase has a complex relationship with plant growth and development, and plays a pivotal role in responding to various biotic and abiotic stresses. The analysis of the U-box gene family in Solanaceae and its expression profile under different stresses holds significant implications. A total of 116 tobacco NtU-boxs and 56 eggplant SmU-boxs were identified based on their respective genome sequences. Phylogenetic analysis of U-box genes in tobacco, eggplant, tomato, Arabidopsis, pepper, and potato revealed five distinct subgroups (IV). Gene structure and protein motifs analysis found a high degree of conservation in both exon/intron organization and protein motifs among tobacco and eggplant U-box genes especially the members within the same subfamily. A total of 15 pairs of segmental duplication and 1 gene pair of tandem duplication were identified in tobacco based on the analysis of gene duplication events, while 10 pairs of segmental duplication in eggplant. It is speculated that segmental duplication events are the primary driver for the expansion of the U-box gene family in both tobacco and eggplant. The promoters of NtU-box and SmU-box genes contained cis-regulatory elements associated with cellular development, phytohormones, environment stress, and photoresponsive elements. Transcriptomic data analysis shows that the expression levels of the tobacco and eggplant U-box genes in different tissues and various abiotic stress conditions. Using cultivar Hongda of tobacco and cultivar Yanzhi of eggplant as materials, qRT-PCR analysis has revealed that 15 selected NtU-box genes and 8 SmU-box may play important roles in response to pathogen Ras invasion both in tobacco and eggplant. [ABSTRACT FROM AUTHOR]

[Objective] Pseudo response regulators (PRRs) are important genes that regulate the flowering pathway in higher plants. The aim of this study is to clone the NtPRR37 gene from Nicotiana tabacum L. and analyze its response to different photoperiods and its effect on flowering, providing a target gene for tobacco flowering regulation. [Method] We cloned NtPRR37 gene from N. tabacum L. by homologous cloning method. Then we performed bioinformatics analysis, tissue-specific expression analysis, and expression pattern analysis of NtPRR37 gene under different light duration treatments. Concurrently, we used Virus Induced Gene Silence (VIGS) to reduce the expression of NtPRR37 and observed the phenotype and detected the expression changes of flowering-related genes. [Result] The fulllength CDS of NtPRR37 was 2 472 bp, encoded a peptide of 823 amino acids with a relative molecular weight of 90.16 kD. It contained REC and CCT domains, which were typical conserved domains of the PRRs gene family. Through homology evolution analysis, it was found that tobacco NtPRR37 belonged to the same branch as PRR37 from Nicotiana tomentosiformis, Nicotiana sylvestris, and Nicotiana benthamiana in evolution. The quantitative reverse transcription-PCR (RT-qPCR) analysis showed that, the expressions of NtPRR37 varied in various tissues of N. tabacum L. at full-bloom stage, with the highest expression in pistils and the lowest expression in lateral roots. Under different photoperiod treatments, the expression of NtPRR37 increased with the prolongation of light exposure. In different light duration treatments, the expression of NtPRR37 increased with the extension of light time, with the lowest expression under darkness, and demonstratted circadian rhythmicity. In NtPRR37-silenced plants, the expression of NtPRR37 was significantly downregulated, and the flowering period was advanced, which may be related to the significant upregulation of flowering-related genes (NtFT4, NtAP1, NtCO, NtSOC1). [Conclusion] The expression of NtPRR37 is regulated by the photoperiod, and NtPRR37 acts as a flowering repressor during the flowering process of tobacco. [ABSTRACT FROM AUTHOR]

Organic amendment and green manuring are two agricultural practices highly recommended to improve sustainability in agriculture since they show numerous beneficial effects on both soils and crops. The main aim of the present study was to evaluate the effect of both, specifically organic fraction municipal solid waste (OFMSW) compost and horse bean (Vicia faba L., cv minor) green manure, combined separately or together with a mineral fertilization using synthetic products and in comparison with a mineral fertilization alone (control), on a top-quality tobacco crop (dark fire-cured Kentucky) grown in the cultivation district of Central Italy (High Tiber Valley, Tuscany region) in 2020 and 2021. The following parameters were measured: (i) leaf emergence rate (LER, leaves day−1); (ii) crop growth rate (CGR, kg dry biomass ha−1 day−1); (iii) root weight density (RWD, mg cm−3); (iv) yield of cured product (CLY, Mg ha−1). Analytical determinations were carried out on soil, sampled at the 0–0.3 m depth (organic matter, %; total N, %; NO3-N, mg kg−1; C/N; P and K, mg kg−1), and on plant biomass (total N, %; NO3-N, kg ha−1). Soil water retention measures were also made. Water productivity (WP, kg cured product m−3 gross crop evapotranspiration, ETc gross), irrigation water use efficiency (IWUE, kg cured product m−3 seasonal irrigation volume) and N agronomic efficiency (NAE, kg cured product kg−1 mineral N applied by synthetic fertilizers) were calculated. Both the applications of OFMSW compost and horse bean green manure increased soil content of organic matter and main nutrients (N, P and K), as well as C/N, when compared with control conditions. There was an increase in soil water content in C conditions over the entire soil matric potential interval (0.04 to 1.2 MPa) with a maximum value at 1.2 MPa in both years. Both practices appeared promising for tobacco cultivation and could help to better address the nitrogen needs of the crop during the season and reduce potential water pollution due to nitrates. Considering the amount of synthetic nitrogen fertilizer saved by using both organic soil amendment and green manuring, there should be fewer potential carbon emissions due to the production, transportation and field application of synthetic nitrogen fertilizers. [ABSTRACT FROM AUTHOR]

Nicotiana tabacum L. (tobacco) has extremely high economic value, medicinal value, scientific research value and some other uses. Though it has been widely cultivated throughout the world, classification and change of its suitable habitats is not that clear, especially in the context of global warming. In order to achieve rational cultivation and sustainable development of tobacco, current (average from 1970-2000) and future (2070, average from 2061-2080) potential suitable habitats of Nicotiana tabacum L. were forecasted with MaxEnt model and ArcGIS platform based on 854 occurrence data and 22 environmental factors in this study. The results revealed that mean temperature of warmest quarter (bio10), annual precipitation (bio12), solar radiation in September (Srad9), and clay content (CLAY) were the four decisive environment variables for the distribution of Nicotiana tabacum L. Under current climate conditions, suitable habitats of Nicotiana tabacum L. were mainly distributed in south-central Europe, south-central North America, most parts of South America, central Africa, south and southeast Asia, and southeast coast of Australia, and only 13.7% of these areas were highly suitable. By the year 2070, suitable habitats under SSP1-2.6, SSP3-7.0, and SSP5-8.5 climate scenarios would all increase with the largest increase found under SSP3-7.0 scenario, while suitable habitats would reduce under SSP2-4.5 climate scenario. Globally, the center of mass of suitable habitats would migrate to southeast to varying degrees within Libya under four different climate scenarios. The emergence of new habitats and the disappearance of old habitats would all occur simultaneously under each climate scenario, and the specific changes in each area, combined with the prediction results under current climate conditions, will provide an important reference for the adjustment of agronomic practices and rational cultivation of Nicotiana tabacum L. both currently and in the future. [ABSTRACT FROM AUTHOR]

Background: The dirigent (DIR) genes encode proteins that act as crucial regulators of plant lignin biosynthesis. In Solanaceae species, members of the DIR gene family are intricately related to plant growth and development, playing a key role in responding to various biotic and abiotic stresses. It will be of great application significance to analyze the DIR gene family and expression profile under various pathogen stresses in Solanaceae species. Results: A total of 57 tobacco NtDIRs and 33 potato StDIRs were identified based on their respective genome sequences. Phylogenetic analysis of DIR genes in tobacco, potato, eggplant and Arabidopsis thaliana revealed three distinct subgroups (DIR-a, DIR-b/d and DIR-e). Gene structure and conserved motif analysis showed that a high degree of conservation in both exon/intron organization and protein motifs among tobacco and potato DIR genes, especially within members of the same subfamily. Total 8 pairs of tandem duplication genes (3 pairs in tobacco, 5 pairs in potato) and 13 pairs of segmental duplication genes (6 pairs in tobacco, 7 pairs in potato) were identified based on the analysis of gene duplication events. Cis-regulatory elements of the DIR promoters participated in hormone response, stress responses, circadian control, endosperm expression, and meristem expression. Transcriptomic data analysis under biotic stress revealed diverse response patterns among DIR gene family members to pathogens, indicating their functional divergence. After 96 h post-inoculation with Ralstonia solanacearum L. (Ras), tobacco seedlings exhibited typical symptoms of tobacco bacterial wilt. The qRT-PCR analysis of 11 selected NtDIR genes displayed differential expression pattern in response to the bacterial pathogen Ras infection. Using line 392278 of potato as material, typical symptoms of potato late blight manifested on the seedling leaves under Phytophthora infestans infection. The qRT-PCR analysis of 5 selected StDIR genes showed up-regulation in response to pathogen infection. Notably, three clustered genes (NtDIR2, NtDIR4, StDIR3) exhibited a robust response to pathogen infection, highlighting their essential roles in disease resistance. Conclusion: The genome-wide identification, evolutionary analysis, and expression profiling of DIR genes in response to various pathogen infection in tobacco and potato have provided valuable insights into the roles of these genes under various stress conditions. Our results could provide a basis for further functional analysis of the DIR gene family under pathogen infection conditions. [ABSTRACT FROM AUTHOR]

ROS-dependent induction of oxidative damage can be used as a trigger initiating genetically determined non-specific protection in plant cells and tissues. Plants are potentially able to withstand various specific (toxic, osmotic) factors of abiotic effects, but do not have sufficient or specific sensitivity to form an adequate effective response. In this work, we demonstrate one of the possible approaches for successful cold acclimation through the formation of effective protection of photosynthetic structures due to the insertion of the heterologous FeSOD gene into the tobacco genome under the control of the constitutive promoter and equipped with a signal sequence targeting the protein to plastid. The increased enzymatic activity of superoxide dismutase in the plastid compartment of transgenic tobacco plants enables them to tolerate the oxidative factor of environmental stresses scavenging ROS. On the other hand, the cost of such resistance is quite high and, when grown under normal conditions, disturbs the arrangement of the intrachloroplastic subdomains leading to the modification of stromal thylakoids, probably significantly affecting the photosynthesis processes that regulate the efficiency of photosystem II. This is partially compensated for by the fact that, at the same time, under normal conditions, the production of peroxide induces the activation of ROS detoxification enzymes. However, a violation of a number of processes, such as the metabolism of accumulation, and utilization and transportation of sugars and starch, is significantly altered, which leads to a shift in metabolic chains. The expected step for further improvement of the applied technology could be both the use of inducible promoters in the expression cassette, and the addition of other genes encoding for hydrogen peroxide-scavenging enzymes in the genetic construct that are downstream in the metabolic chain. [ABSTRACT FROM AUTHOR]

Southwestern China is receiving excessive chemical fertilizers to meet the challenges of continuous cropping. These practices are deteriorating the soil environment and affecting tobacco (Nicotiana tabacum L.) yield and quality adversely. A novel microbially enriched biochar-based fertilizer was synthesized using effective microorganisms, tobacco stalk biochar and basal fertilizer. A field-scale study was conducted to evaluate the yield response of tobacco grown on degraded soil amended with our novel biochar-based microbial fertilizer (BF). Four treatments of BF (0%, 1.5%, 2.5% and 5%) were applied in the contaminated field to grow tobacco. The application of BF1.5, BF2.5 and BF5.0 increased the available water contents by 9.47%, 1.18% and 2.19% compared to that with BF0 respectively. Maximum growth of tobacco in terms of plant height and leaf area was recorded for BF1.5 compared to BF0. BF1.5, BF2.5 and BF5.0 increased SPAD by 13.18–40.53%, net photosynthetic rate by 5.44–60.42%, stomatal conductance by 8.33–44.44%, instantaneous water use efficiency by 55.41–93.24% and intrinsic water use efficiency by 0.09–24.11%, while they decreased the intercellular CO2 concentration and transpiration rate by 3.85–6.84% and 0.29–47.18% relative to BF0, respectively (p < 0.05). The maximum increase in tobacco yield was recorded with BF1.5 (23.81%) compared to that with BF0. The present study concludes that the application of BF1.5 improves and restores the degraded soil by improving the hydraulic conductivity and by increasing the tobacco yield. [ABSTRACT FROM AUTHOR]

Two new isoindolin-1-ones (1 and 2), were isolated from the stems of Nicotiana tabacum (a cultivar of sun-cured tobacco with high resistance to black shank diseases, which originated from Huize local sun-cured tobacco by ethyl methanesulfonate mutagenesis). Their structures were elucidated using spectroscopic methods, including extensive 1D and 2D NMR techniques. Interestingly, both compounds showed high antibacterial activity with bacteriostatic diameter within the range 18.2 ± 1.4–25.8 ± 2.2 mm against five pathogenic strains, and they have the potential values for use as an antiseptic agent for Heat Not Burning cigarette-tipping paper. [ABSTRACT FROM AUTHOR]

Soil drought is among the serious problems in agriculture leading to considerable losses of crop production. Tobacco is important industrial crop, which is relatively sensitive to water deficit, and the selection of drought resistant varieties is of particular interest. In line with this, we examined the drought tolerance of three genotypes of flue-cured Virginia tobacco-cv. Coker 254 (American origin), line L 825 (Czech origin), which are introduced in Bulgaria, and their hybrid cv. Virzhinia 0514, which is certified as Bulgarian standard. The content of stress biomarkers malondialdehyde, hydrogen peroxide and free proline as well as some morphometric traits were assessed in plants subjected to three levels of drought. The results showed that the three tested tobacco genotypes were moderately tolerant to drought. These data can be used for selection and development of new genotypes with improved tolerance to soil drought. [ABSTRACT FROM AUTHOR]

The present study aimed to test a multipurpose sustainable tobacco farming system allowing more efficient use of production factors (e.g., mineral N fertilizer) thanks to larger commercial yields, albeit diversified (smoke products, bioactive compounds for nutraceutical and cosmeceutical uses, energy), per unit of land area. Three tobacco types (dark air-cured, IBG; light air-cured, Bu; dark fire-cured, Ky) were grown in the field in 2021 on three different soils (sandy clay loam, SCL; sandy loam, SL; clay loam, CL). The total waste biomass (WB, kg dry weight, d.w. ha−1) was measured. Commercial leaves yield (CLY, kg d.w. ha−1), N agronomic efficiency (NAE, kg d.w. kg−1 N), total polyphenols content (TP, mg kg−1 d.w.), antioxidant activity (ABTS, DPPH and FRAP, mmol Trolox Equivalent, TE, kg−1 d.w.) and yield of polyphenols (PY, kg ha−1) were determined. The calorific value (CV, MJ kg−1 d.w.), volatile matter (VM, %) and ash contents (%) were also measured, and biomass energy yield (BEY, GJ ha−1 yr−1) was then calculated. Very high percentages (>40%) of total biomass produced by the different tobacco types were pre-harvest waste. NAE increased by 2- to more than 8-fold thanks to a greater potential commercial biomass produced with the same amount of N fertilizer used. Four main components were found in the tobacco polyphenols profile, namely 3-O-CQA, luteolin 7 rutinoside, rutin and quinic acid, which accounted for more than 80% of TP. BEY ranged between 122.3 GJ ha−1 yr−1 (Bu) and 29.9 GJ ha−1 yr−1 (Ky). Both polyphenols yield and energy potential per unit land area and/or per growing season appeared competitive with those from other herbaceous crops. The proposed multipurpose system appeared as a production circuit characterized by a virtuous and sustainable flow of resources. [ABSTRACT FROM AUTHOR]

Sub-group 4 R2R3-type MYB transcription factors (TFs) are involved in repression of the phenylpropanoid pathway. In this study, we identified and characterized three MYB transcriptional repressors in common tobacco, named NtMYB1, NtMYB2, and NtMYB4, which belong to sub-group 4 members. Tissue expression properties and specificity of stress-induced expression indicated that the NtMYB1, NtMYB2, and NtMYB4 may perform different functions in tobacco. Overexpression of NtMYB1, NtMYB2, and NtMYB4 reduced phenylpropanoids and flaonoid compounds, such as caffeic acid, chlorogenic acid, neochlorogenic acid, and rutin, respectively. Meanwhile, the expression of the most structural genes involved in phenylpropanoids and flavonoids were reduced in the overexpression transgenic lines. In addition, results based on co-expression and expression analyses revealed possible interactions between NtMYB1, NtMYB2, and NtMYB4 and multiple bHLH TFs. Our results showed that the three sub-group 4 R2R3 members exhibited different expression pattern in various tissues and under abiotic stress conditions. Meanwhile, we hypothesized that the three of them involved in the inhibition of phenylpropanoid and flavonoid biosynthesis might be interaction with bHLH TFs.

The aim of the work was to investigate detection of different modifications of the green fluorescent protein gene (gfp) in the transgenic tobacco and maize plants by polymerase chain reaction (PCR). Methods. Total DNA isolation, PCR, electrophoresis of DNA in agarose gel, bioinformatic resources. Results. Three pairs of primers were used for PCR analysis of tobacco and maize containing wild-type gfp or mutant synthetic gene S65Tpgfp. The primer pair gfp1F-gfp1R interacted with the wild-type gfp gene only. The gfp2F-gfp2R primers interacted with the gfp gene of different modifications both in tobacco and maize. The gfp3F-gfp3R primer pair interacted with the modified S65Tpgfp gene in tobacco DNA, but not with maize samples. Conclusions. Primers for detection of heterologous gfp gene, which were both narrowly specific (only one gene modification could be detected), and universal (more than one gene modification could be detected), were verified. It was shown that the primer pair gfp2F-gfp2R was universal for gfp gene detection both in tobacco and maize plants by PCR. The results obtained with gfp2F-gfp2R were reliably reproducible, so this primer pair is recommended for general use.

Abstract Background Glutathione S-transferases (GSTs) are large and multifunctional proteases that play an important role in detoxification, protection against biotic and abiotic stresses, and secondary metabolite transportation which is essential for plant growth and development. However, there is limited research on the identification and function of NtGSTs. Results This study uses K326 and other six tobacco varieties (Hongda, HG, GDH11, Va116, VG, and GDH88) as materials to conduct comprehensive genome-wide identification and functional characterization of the GST gene in tobacco. A total of 59 NtGSTs were identified and classified into seven subfamilies via the whole-genome sequence analysis, with the Tau type serving as the major subfamily. The NtGSTs in the same branch of the evolutionary tree had similar exon/intron structure and motif constitution. There were more than 42 collinear blocks between tobacco and pepper, tomato, and potato, indicating high homology conservation between them. Twelve segmental duplicated gene pairs and one tandem duplication may have had a substantial impact on the evolution and expansion of the tobacco GST gene family. The RT-qPCR results showed that the expression patterns of NtGSTs varied significantly among tissues, varieties, and multiple abiotic stresses, suggesting that NtGST genes may widely respond to various abiotic stresses and hormones in tobacco, including NtGSTF4, NtGSTL1, NtGSTZ1, and NtGSTU40. Conclusions This study provides a comprehensive analysis of the NtGST gene family, including structures and functions. Many NtGSTs play a critical regulatory role in tobacco growth and development, and responses to abiotic stresses. These findings offer novel and valuable insights for understanding the biological function of NtGSTs and the reference materials for cultivating highly resistant varieties and enhancing the yield and quality of crops.

Metabolic changes under stress are often studied in short-term experiments, revealing rapid responses in gene expression, enzyme activity, and the amount of antioxidants. In a long-term experiment, it is possible to identify adaptive changes in both primary and secondary metabolism. In this study, we characterized the physiological state of tobacco plants and assessed the amount and spectrum of phenolic compounds and the lignification of axial organs under excess copper stress in a long-term experiment (40 days). Plants were treated with 100 and 300 μM CuSO4, as well as a control (Knop solution). Copper accumulation, the size and anatomical structure of organs, stress markers, and the activity of antioxidant enzymes were studied. Lignin content was determined with the cysteine-assisted sulfuric method (CASA), and the metabolite profile and phenolic spectrum were determined with UHPLC-MS and thin-layer chromatography (TLC). Cu2+ mainly accumulated in the roots and, to a lesser extent, in the shoots. Copper sulfate (100 μM) slightly stimulated stem and leaf growth. A higher concentration (300 μM) caused oxidative stress; H2O2 content, superoxide dismutase (SOD), and guaiacol peroxidase (GPOX) activity increased in roots, and malondialdehyde (MDA) increased in all organs. The deposition of lignin increased in the roots and stems compared with the control. The content of free phenolics, which could be used as substrates for lignification, declined. The proportions of ferulic, cinnamic, and p-coumaric acids in the hydrolysate of bound phenolics were higher, and they tended toward additional lignification. The metabolic profile changed in both roots and stems at both concentrations, and changed in leaves only at a concentration of 300 μM. Thus, changes in the phenolic spectrum and the enhanced lignification of cell walls in the metaxylem of axial (root and stem) organs in tobacco can be considered important metabolic responses to stress caused by excess CuSO4. [ABSTRACT FROM AUTHOR]

ROS-dependent induction of oxidative damage can be used as a trigger initiating genetically determined non-specific protection in plant cells and tissues. Plants are potentially able to withstand various specific (toxic, osmotic) factors of abiotic effects, but do not have sufficient or specific sensitivity to form an adequate effective response. In this work, we demonstrate one of the possible approaches for successful cold acclimation through the formation of effective protection of photosynthetic structures due to the insertion of the heterologous FeSOD gene into the tobacco genome under the control of the constitutive promoter and equipped with a signal sequence targeting the protein to plastid. The increased enzymatic activity of superoxide dismutase in the plastid compartment of transgenic tobacco plants enables them to tolerate the oxidative factor of environmental stresses scavenging ROS. On the other hand, the cost of such resistance is quite high and, when grown under normal conditions, disturbs the arrangement of the intrachloroplastic subdomains leading to the modification of stromal thylakoids, probably significantly affecting the photosynthesis processes that regulate the efficiency of photosystem II. This is partially compensated for by the fact that, at the same time, under normal conditions, the production of peroxide induces the activation of ROS detoxification enzymes. However, a violation of a number of processes, such as the metabolism of accumulation, and utilization and transportation of sugars and starch, is significantly altered, which leads to a shift in metabolic chains. The expected step for further improvement of the applied technology could be both the use of inducible promoters in the expression cassette, and the addition of other genes encoding for hydrogen peroxide-scavenging enzymes in the genetic construct that are downstream in the metabolic chain.

Southwestern China is receiving excessive chemical fertilizers to meet the challenges of continuous cropping. These practices are deteriorating the soil environment and affecting tobacco (Nicotiana tabacum L.) yield and quality adversely. A novel microbially enriched biochar-based fertilizer was synthesized using effective microorganisms, tobacco stalk biochar and basal fertilizer. A field-scale study was conducted to evaluate the yield response of tobacco grown on degraded soil amended with our novel biochar-based microbial fertilizer (BF). Four treatments of BF (0%, 1.5%, 2.5% and 5%) were applied in the contaminated field to grow tobacco. The application of BF1.5, BF2.5 and BF5.0 increased the available water contents by 9.47%, 1.18% and 2.19% compared to that with BF0 respectively. Maximum growth of tobacco in terms of plant height and leaf area was recorded for BF1.5 compared to BF0. BF1.5, BF2.5 and BF5.0 increased SPAD by 13.18–40.53%, net photosynthetic rate by 5.44–60.42%, stomatal conductance by 8.33–44.44%, instantaneous water use efficiency by 55.41–93.24% and intrinsic water use efficiency by 0.09–24.11%, while they decreased the intercellular CO2 concentration and transpiration rate by 3.85–6.84% and 0.29–47.18% relative to BF0, respectively (p < 0.05). The maximum increase in tobacco yield was recorded with BF1.5 (23.81%) compared to that with BF0. The present study concludes that the application of BF1.5 improves and restores the degraded soil by improving the hydraulic conductivity and by increasing the tobacco yield.

Two new aromatic sesquiterpenes, 1-isopropyl-5-methoxy-3,7-dimethylnaphthalene (1) and 3-hydroxymethyl-1-isopropyl-5-methoxy-7-methylnaphthalene (2), as well as five known sesquiterpenes (3–7), were isolated from the leaves of Nicotiana tabacum (a mutant tobacco for solanesol synthesis gene knockout by CRISPR-Cas9 gene editing). Their structures were determined by spectroscopic methods, including extensive 1D and 2D NMR techniques. Compounds 1 and 2 were tested for their anti-methicillin-resistant Staphylococcus aureus (anti-MRSA) activity. The results revealed that compounds 1 and 2 showed good inhibition with IZD of 12.8 ± 2.0 and 15.2 ± 2.2 mm, respectively. In addition, compounds 1 and 2 have a weak floral fragrance, and they have the potential for use as an essence for cigarettes. [ABSTRACT FROM AUTHOR]

Copyright of Academic Journal of Agriculture / Akademik Ziraat Dergisi is the property of University of Ordu, Faculty of Agriculture, Academic Journal of Agriculture and its content may not be copied or emailed to multiple sites or posted to a listserv without the copyright holder's express written permission. However, users may print, download, or email articles for individual use. This abstract may be abridged. No warranty is given about the accuracy of the copy. Users should refer to the original published version of the material for the full abstract. (Copyright applies to all Abstracts.)

Tobacco (Nicotiana tabacum L.) is an important industrial crop grown in more than one hundred countries worldwide, with high adaptability on a great variety of soils, and under quite diverse climatic conditions. Information regarding the chemical composition of tobacco leaves is important for the tobacco industry because it generally may exhibit important differences among different locations. In this study, five oriental and six flue and air-cured type tobacco varieties were cultivated in five areas of Greece for three cultivation periods, in order to evaluate the effect of these three factors (variety, area, and period), on the yield as well as nicotine content, and sugar and nitrate content of the produced leaves. For the oriental type varieties, the results showed that the area exhibited significant interaction with both variety and the cultivation year, related to the yield. Concerning the nicotine content, no significant differences were observed regarding the areas of experimentation; however, it significantly varied within the varieties. In particular, the nicotine content of Katerini 53 exhibited a significant difference compared to Xanthi 2A, which had the highest content of all tested varieties. Sugar content was affected significantly by all the factors considered, with the area of Xanthi exhibiting lower values compared to all other varieties. Regarding the flue and air-cured varieties, the only interaction that was found to be statistically significant, when the yield was assessed, was between the cultivation year and the area. It was found that there was a significant interaction between cultivation year and area for the nicotine content as well. Furthermore, cultivation year and variety significantly affected the sugar content. The varieties Burley 21E and NC7LC gave the lowest sugars. In general, it was observed that in the case of the oriental type varieties, the variety adaptation in specific regions resulted in higher yields, with the paradigmatic example of the variety Katerini 53 in the Katerini area, and Xanthi 81 in the Xanthi area. On the other hand, in the case of the flue and air-cured varieties, Karditsa was the area that exhibited the lowest yield of all tested varieties. [ABSTRACT FROM AUTHOR]

Background: Glutathione S-transferases (GSTs) are large and multifunctional proteases that play an important role in detoxification, protection against biotic and abiotic stresses, and secondary metabolite transportation which is essential for plant growth and development. However, there is limited research on the identification and function of NtGSTs. Results: This study uses K326 and other six tobacco varieties (Hongda, HG, GDH11, Va116, VG, and GDH88) as materials to conduct comprehensive genome-wide identification and functional characterization of the GST gene in tobacco. A total of 59 NtGSTs were identified and classified into seven subfamilies via the whole-genome sequence analysis, with the Tau type serving as the major subfamily. The NtGSTs in the same branch of the evolutionary tree had similar exon/intron structure and motif constitution. There were more than 42 collinear blocks between tobacco and pepper, tomato, and potato, indicating high homology conservation between them. Twelve segmental duplicated gene pairs and one tandem duplication may have had a substantial impact on the evolution and expansion of the tobacco GST gene family. The RT-qPCR results showed that the expression patterns of NtGSTs varied significantly among tissues, varieties, and multiple abiotic stresses, suggesting that NtGST genes may widely respond to various abiotic stresses and hormones in tobacco, including NtGSTF4, NtGSTL1, NtGSTZ1, and NtGSTU40. Conclusions: This study provides a comprehensive analysis of the NtGST gene family, including structures and functions. Many NtGSTs play a critical regulatory role in tobacco growth and development, and responses to abiotic stresses. These findings offer novel and valuable insights for understanding the biological function of NtGSTs and the reference materials for cultivating highly resistant varieties and enhancing the yield and quality of crops. [ABSTRACT FROM AUTHOR]

Concerning tobacco (Nicotiana tabacum L.) producers the size and color of the leaf are important factors in understanding the quality grade of tobacco leaves in the market. The color of tobacco leaves serves as an indicator of quality and is referred to as the maturity index when determining the optimal time for harvesting. In this study, a hybrid mobile application was developed to help determine the harvest time. CoLab was preferred as the backend. Python Imaging Library (Pillow) was used for image processing on the server side. Color correction was performed on the images taken with the help of X-rite. The correlated color temperature (CCT) value of the repaired images was calculated. The CCT values were calculated using the Ohno method. Quality grade (QG) was calculated using the mean CCT value. The data of the images obtained depending on the time were used in the application as a graphic. We present quality grade of tobacco automatically identifying the plant leaves in a given image with the help of the mobile application. [ABSTRACT FROM AUTHOR]

Copyright of Journal of Agricultural Sciences, Belgrade is the property of University of Belgrade, Faculty of Agriculture and its content may not be copied or emailed to multiple sites or posted to a listserv without the copyright holder's express written permission. However, users may print, download, or email articles for individual use. This abstract may be abridged. No warranty is given about the accuracy of the copy. Users should refer to the original published version of the material for the full abstract. (Copyright applies to all Abstracts.)

Tobacco has a strong cadmium (Cd) enrichment capacity, meaning that it can absorb large quantities from the environment, but too much Cd will cause damage to the plant. It is not yet clear how the plant can dynamically respond to Cd stress. Here, we performed a temporal transcriptome analysis of tobacco roots under Cd treatment from 0 to 48 h. The number of differentially expressed genes (DEGs) was found to change significantly at 3 h of Cd treatment, which we used to define the early and middle stages of the Cd stress response. The gene ontology (GO) term analysis indicates that genes related to photosynthesis and fatty acid synthesis were enriched during the early phases of the stress response, and in the middle phase biological process related to metal ion transport, DNA damage repair, and metabolism were enriched. It was also found that plants use precursor mRNA (pre-mRNA) processes to first resist Cd stress, and with the increasing of Cd treatment time, the overlapped genes number of DEGs and DAS increased, suggesting the transcriptional levels and post-transcriptional level might influence each other. This study allowed us to better understand how plants dynamically respond to cadmium stress at the transcriptional and posttranscriptional levels and provided a reference for the screening of Cd-tolerant genes in the future. [ABSTRACT FROM AUTHOR]

Catalase (CAT) is an enzyme encoded by the catalase gene family that plays an important role in the removal of reactive oxygen species. In this study, seven CAT genes were identified in Nicotiana tabacum L. and were classified into three groups. Gene structure analysis revealed that NtCAT1–6 has six or seven introns while NtCAT7 only contains one. The relative position of introns in NtCAT1 and NtCAT2 had high similarity. Tissue-specific analysis shows that NtCAT1–4 were expressed intensively in the shoot while NtCAT5 and NtCAT6 were in the root. NtCAT7 expression was influenced by circadian rhythms. NtCATs expression had the greatest change under drought stress. Additionally, expression of NtCAT5, NtCAT6 and NtCAT7 were upregulated under cold stress but downregulated under drought and salt stress. This study will help in understanding the behavior of CAT genes during environmental stress in tobacco. [ABSTRACT FROM AUTHOR]

Industrial wastewater obtained from hydrothermal liquefaction (HTL-WW) of food wastes for biofuels production could represent a source of crop nutrients since it is characterized by a high amount of organic and inorganic compounds. In the present work, the potential use of HTL-WW as irrigation water for industrial crops was investigated. The composition of the HTL-WW was rich in nitrogen, phosphorus, and potassium with high level of organic carbon. A pot experiment with Nicotiana tabacum L. plants was conducted using diluted wastewater to reduce the concentration of some chemical elements below the official accepted threshold values. Plants were grown in the greenhouse under controlled conditions for 21 days and irrigated with diluted HTL-WW every 24 hours. Soils and plants were sampled every seven days to evaluate, over time, the effect of wastewater irrigation both on soil microbial populations, through high-throughput sequencing, and plant growth parameters, through the measurement of different biometric indices. Metagenomic results highlighted that, in the HTL-WW treated rhizosphere, the microbial populations shifted via their mechanisms of adaptation to the new environmental conditions, establishing a new balance among bacterial and fungal communities. Identification of microbial taxa occurring in the rhizosphere of tobacco plants during the experiment highlighted that the HTL-WW application improved the growth of Micrococcaceae, Nocardiaceae and Nectriaceae, which included key species for denitrification, organic compounds degradation and plant growth promotion. As a result, irrigation with HTL-WW improved the overall performance of tobacco plants which showed higher leaf greenness and increased number of flowers compared to irrigated control plants. Overall, these results demonstrate the potential feasibility of using of HTL-WW in irrigated agriculture.

We investigated the contribution of two Arabidopsis AtHIRD11 (AT1G54410, DH1) and AtLTI30 (AT3G50970, DH3) dehydrin genes to cadmium stress tolerance in plants. We generated two sets of transgenic tobacco plants overexpressing dehydrins DH1 and DH3. Transgenic and non-transgenic (control) plants were exposed to the long-term (50 µM cadmium chloride for 10 days) and short-term (100 µM cadmium chloride for 24 h) stress and analysed. Our results revealed that the dehydrin DH3 has modulated the cadmium uptake capacity of tobacco. The DH3 overexpressing plants absorbed cadmium significantly faster and in larger quantitates (at least 2.4-fold). Most of the absorbed cadmium was retained in the roots and not translocated to the leaves of DH3 plants. In root cells, a larger portion of cadmium was allocated in the soluble fraction consisting of cytosol and vacuoles, suggesting a potential role for DH3 in cadmium detoxification. An overexpression of the gene DH3 led to a significant (at least 6- and 2.6-fold) increase in the content of non-enzymatic antioxidants chlorogenic acid and rutin, respectively, under standard as well as stress conditions. Some of the cadmium-responsive miRNAs (nta-miR169q, nta-miR319b and nta-miR156g) were supressed in DH3 overexpressing plants, indicating a potential role for DH3 in modulating of plant's defence response. Unlike the DH3 gene, the contribution of the dehydrin DH1 to cadmium tolerance seems to be insignificant. [ABSTRACT FROM AUTHOR]

Summary: The most important factor in tobacco is quality. It is the sum of physical, chemical, and smoking properties that characterize quality. Post‐harvest curing of leaves plays an important role in quality. In this study, an approach based on correlated color temperature (CCT) is proposed to determine the ideal curing time of tobacco leaves. While photographing leaf samples from different days during the curing of tobacco leaves, color correction was made using X‐rite color checker to eliminate the negative effect of ambient light. CCT values were calculated from the repaired photographs using the McCammy method. The inclusion rate of pixels during the calculation is 97%. The equation of CCT variation depending on the day is fitted as CCT=(−0.1889×day3)+(4.8347×day2)−(115.59×day)+4919.2$$ \mathrm{CCT}=\left(-0.1889\times {\mathrm{day}}^3\right)+\left(4.8347\times {\mathrm{day}}^2\right)-\left(115.59\times \mathrm{day}\right)+4919.2 $$. RMSE (26.77 K) and high R2$$ {R}^2 $$ value were calculated as 98.8% in the equation. CCT values have been converted to quality grade (QG) values to be more understandable. As a result of the study, CCT values ranged between 5000 and 2500 K and corresponding QG values ranged between 79% and 105%. The ideal curing time was calculated as the 17th day after harvesting, the QG value for that day was 100% and the average CCT value was 3000 K. [ABSTRACT FROM AUTHOR]

The mode of inheritance and heterotic effect were studied in ten F1 crosses obtained by one-way diallel crossing between five parental genotypes: MV-1, P 76/86, Adiyaman, Basma-Djebel and P 66 9 7. The following quantitative traits were studied: the number of leaves per stalk, length of leaves from the middle belt of the stalk and yield of green leaf mass per stalk and per hectare. The trial was set up in the experimental field of Scientific Tobacco Institute-Prilep, using a randomized block design with four replications in the period 2018- 2019. Traditional cultural practices were applied during the growing season of tobacco in the field. The aim of this research was to study the mode of inheritance of the quantitative traits, to detect heterosis in the F1 generation and to assess its economic viability. Analysis of variance determined statistically significant differences in traits between parents and hybrids in the two-year investigation. The most common way of trait inheritance is partial-dominant, then intermediate. The negative heterotic effect on the number of leaves per stalk has P 76/86 x P 66 9 7. The hybrids MV-1 x Adiyaman, P 76/86 x Basma-Djebel, P 76/86 x P 66 9 7 and Basma-Djebel x P 66 9 7 have a positive heterotic effect on the length of the leaves. Oriental hybrids, where one of the parents is the variety P 66 9 7, have positive heterosis in the yield of green leaf mass per stalk. The investigation provides very useful guidance for future successive selection activities. [ABSTRACT FROM AUTHOR]

The SQUAMOSA promoter binding protein-like (SPL)SPL family genes play an important role in regulating plant growth and development, synthesis of secondary metabolites, and resistance to stress. Understanding of the role of the SPL family in tobacco is still limited. In this study, 42 NtSPL genes were identified from the genome of the tobacco variety TN90. According to the results of the conserved motif and phylogenetic tree, the NtSPL genes were divided into eight subgroups, and the genes in the same subgroup showed similar gene structures and conserved domains. The cis-acting element analysis of the NtSPL promoters showed that the NtSPL genes were regulated by plant hormones and stresses. Twenty-eight of the 42 NtSPL genes can be targeted by miR156. Transcriptome data and qPCR results indicated that the expression pattern of miR156-targeted NtSPL genes was usually tissue specific. The expression level of miR156 in tobacco was induced by Cd stress, and the expression pattern of NtSPL4a showed a significant negative correlation with that of miR156. These results suggest that miR156-NtSPL4a may mediate the tobacco response to Cd stress. This study lays a foundation for further research on the function of the NtSPL gene and provides new insights into the involvement of NtSPL genes in the plant response to heavy metal stress. [ABSTRACT FROM AUTHOR]

This work is focused on the production of transgenic tobacco plants overexpressing the oak dehydrin gene AY607707.1. Transgenic tobacco plants were generated via Agrobacterium-mediated transformation. The T-DNA of the plant transformation vector pMK contained the dehydrin gene fused to the constitutive double dCaMV35S promoter and the selectable neomycin phosphotransferase gene. The sequence of the dehydrin gene was isolated from Quercus robur by PCR approach and cloned. The constructed binary vector pMK was introduced into Agrobacterium tumefaciens LBA4404 and used in the transformation experiments. Transgenic plants were generated with an efficiency of 32.7%. PCR analyses confirmed the transgenic nature of regenerated T0 plants. The expression of the oak dehydrin gene was proved by RT-PCR and quantified by qPCR analyses. [ABSTRACT FROM AUTHOR]

[Display omitted] • Transcriptional mechanism of tobacco seed deterioration was revealed by small RNA and transcriptome sequencing. • The cleavage relationship between tobacco miRNA and mRNA was validated by degradome sequencing. • Coherent miRNA-mRNA pairs of seed deterioration were screened through combined analysis. • Rice homologous target genes found that OsARF18 and OsMBD707 were functional in seed deterioration. • Transcriptional multiomics provides substantial transcriptional data regarding seed deterioration for further study. Mature seeds deteriorate gradually and die eventually during long-term storage. Controlled deterioration is often used to accelerate the seed deterioration rate to assess the seed vigor and physiological quality of seed lots. Although it is well known that the process of seed deterioration produced by controlled deterioration is distinct from that caused by long-term storage, the differences in transcriptional levels have not been reported. Clarifying the mechanism of seed deterioration is critical for identifying, conserving and utilizing germplasm resources. Tobacco (Nicotiana tabacum L.) seeds were studied thoroughly using transcriptome, small RNA, and degradome sequencing after long-term storage (LS) and controlled deterioration (CD). Co-expression trend analysis identified transcripts involved in tobacco seed deterioration, while phylogenetic analysis helped to uncover comparable targets in rice (Oryza sativa L.) for further verification and utilization. In LS and CD, a total of 2,112 genes and 164 miRNAs were differentially expressed, including 20 interaction miRNA-mRNA pairs with contrasting expression. Transcriptional multiomics found that the main causes of LS were plant hormone signal transduction and protein processing in the endoplasmic reticulum, whereas the primary cause of CD was nucleotide excision repair dysfunction. The homeostatic balance of RNA degradation and the spliceosome occurred in both modes of seed deterioration. Additionally, co-expression trend analysis identified two coherent pairs, nta-miR160b - NtARF18 and nta-miR396c - NtMBD10 , as being significant in LS and CD, respectively. For utilization, rice homologous targets OsARF18 and OsMBD707 were verified to play similar roles in LS and CD, respectively. This study demonstrated the transcriptional mechanism of tobacco and key genes in seed deterioration. And the application of key genes in rice also verified the feasibility of the multiomics method, guiding the identification of candidate genes to precisely delay seed deterioration in other species of seed research. [ABSTRACT FROM AUTHOR]

Abstract Background The basic leucine zipper (bZIP) transcription factor (TF) is one of the largest families of transcription factors (TFs). It is widely distributed and highly conserved in animals, plants, and microorganisms. Previous studies have shown that the bZIP TF family is involved in plant growth, development, and stress responses. The bZIP family has been studied in many plants; however, there is little research on the bZIP gene family in tobacco. Results In this study, 77 bZIPs were identified in tobacco and named NtbZIP01 through to NtbZIP77. These 77 genes were then divided into eleven subfamilies according to their homology with Arabidopsis thaliana. NtbZIPs were unevenly distributed across twenty-two tobacco chromosomes, and we found sixteen pairs of segmental duplication. We further studied the collinearity between these genes and related genes of six other species. Quantitative real-time polymerase chain reaction analysis identified that expression patterns of bZIPs differed, including in different organs and under various abiotic stresses. NtbZIP49 might be important in the development of flowers and fruits; NtbZIP18 might be an important regulator in abiotic stress. Conclusions In this study, the structures and functions of the bZIP family in tobacco were systematically explored. Many bZIPs may play vital roles in the regulation of organ development, growth, and responses to abiotic stresses. This research has great significance for the functional characterisation of the tobacco bZIP family and our understanding of the bZIP family in higher plants.

Tobacco is one of the important cash crops in China. It is sensitive to low temperature, especially in the early stage of tobacco seed germination and seedling growth. Low temperature stress directly affects the germination of tobacco seeds, leading to irregular emergence and slow growth of seedlings. Therefore, it is important to improve the vigor and cold tolerance of tobacco. In this study, two cold-sensitive tobacco varieties were primed individually with ZnSO 4 , FeSO 4 and Na 2 SeO 3 to test the effects of different microelements priming on seed germination. The results showed that under low temperature (11 °C), all three elements could improve tobacco seed vigor and promote seed germination, with selenium priming exhibiting the best effect. Selenium priming significantly increased the seed vigor index of YY97 by 29.60 % and of YY85 by 47.57 %. Moreover, selenium priming could effectively enhance POD activity in seeds, promote the oxidation of phenolic substances and enhance the activities of G-6-PDH and 6-P-GDH in the process of scavenging ROS and converting H 2 O 2 to ·O 2 −. The results suggested that selenium played an important role in promoting tobacco seed germination under low temperature. • Zn, Fe and Se priming improved tobacco seed germination and seedling establishment at low temperature. • Se priming exhibited the strongest promotion effect on the anti-oxidation system. • POD activity was particularly enhanced by Se priming. • The increased POD activity promoted seed germination from multiple physiological aspects. [ABSTRACT FROM AUTHOR]

Metabolic changes under stress are often studied in short-term experiments, revealing rapid responses in gene expression, enzyme activity, and the amount of antioxidants. In a long-term experiment, it is possible to identify adaptive changes in both primary and secondary metabolism. In this study, we characterized the physiological state of tobacco plants and assessed the amount and spectrum of phenolic compounds and the lignification of axial organs under excess copper stress in a long-term experiment (40 days). Plants were treated with 100 and 300 μM CuSO4, as well as a control (Knop solution). Copper accumulation, the size and anatomical structure of organs, stress markers, and the activity of antioxidant enzymes were studied. Lignin content was determined with the cysteine-assisted sulfuric method (CASA), and the metabolite profile and phenolic spectrum were determined with UHPLC-MS and thin-layer chromatography (TLC). Cu2+ mainly accumulated in the roots and, to a lesser extent, in the shoots. Copper sulfate (100 μM) slightly stimulated stem and leaf growth. A higher concentration (300 μM) caused oxidative stress; H2O2 content, superoxide dismutase (SOD), and guaiacol peroxidase (GPOX) activity increased in roots, and malondialdehyde (MDA) increased in all organs. The deposition of lignin increased in the roots and stems compared with the control. The content of free phenolics, which could be used as substrates for lignification, declined. The proportions of ferulic, cinnamic, and p-coumaric acids in the hydrolysate of bound phenolics were higher, and they tended toward additional lignification. The metabolic profile changed in both roots and stems at both concentrations, and changed in leaves only at a concentration of 300 μM. Thus, changes in the phenolic spectrum and the enhanced lignification of cell walls in the metaxylem of axial (root and stem) organs in tobacco can be considered important metabolic responses to stress caused by excess CuSO4.

Drought stress usually causes huge economic losses for tobacco industries. Drought stress exhibits multifaceted impacts on tobacco systems through inducing changes at different levels, such as physiological and chemical changes, changes of gene transcription and metabolic changes. Understanding how plants respond and adapt to drought stress helps generate engineered plants with enhanced drought resistance. In this study, we conducted multiple time point-related physiological, biochemical,transcriptomic and metabolic assays using K326 and its derived mutant 28 (M28) with contrasting drought tolerance. Through integrative analyses of transcriptome and metabolome,we observed dramatic changes of gene expression and metabolic profiles between M28 and K326 before and after drought treatment. we found that some of DEGs function as key enzymes responsible for ABA biosynthesis and metabolic pathway, thereby mitigating impairment of drought stress through ABA signaling dependent pathways. Four DEGs were involved in nitrogen metabolism, leading to synthesis of glutamate (Glu) starting from NO−3 /NO−2 that serves as an indicator for stress responses. Importantly, through regulatory network analyses, we detected several drought induced TFs that regulate expression of genes responsible for ABA biosynthesis through network, indicating direct and indirect involvement of TFs in drought responses in tobacco. Thus, our study sheds some mechanistic insights into how plant responding to drought stress through transcriptomic and metabolic changes in tobacco. It also provides some key TF or non-TF gene candidates for engineering manipulation for breeding new tobacco varieties with enhanced drought tolerance.

To screen the upstream regulatory transcription factors of cembratrienol synthase（NtCBT）in the cembranoids synthesis pathway of tobacco（Nicotiana tabacum L.），the promoter of NtCBT gene was cut into P1 to P6 regions，six yeast one-hybrid bait vectors pAbAi-Px were constructed and transformed into Y1H competent yeast cells，bait strains were constructed，and a self-activation experiment was completed. Furthermore，a screening was finished from yeast cDNA library of tobacco trichomes in order to obtain the transcription factors that interacted with P5 regions（-279 - -119 bp）. The results showed that，the growths of the strains containing P1 to P5 regions bait strain were inhibited on the medium added with 200 ng/mL AbA. A total of 49 positive colonies were obtained through Y1H screening when P5 regions as bait strain，among them，35 colonies were non-repeating sequences，and 3 of colonies were annotated as transcription factors of ANL2， ML1 and NF-Y. Above results lay a foundation for further study of gene expression and regulation mechanism of NtCBT. [ABSTRACT FROM AUTHOR]

This study aimed to compare recombinant inbred lines (RIL) obtained via the bulk method with doubled haploid lines (DHL). For comparison, 190 DHL and 194 RIL were evaluated simultaneously at two different locations. Phenotypic and genetic parameters were estimated for the traits green leaf yield (GLY) and alkaloid content (ALK). The results of the RIL and DHL evaluations were similar for ALK. Conversely, estimates of genetic variation and heritability in the DHL were higher than those in the RIL for GLY. However, the mean estimate of the RIL was 13.3% higher than that of the DHL and thus, the annual gain with selection was higher for the RIL. The use of DHL in a breeding program program will be efficient in comparison to RIL if a large number of lines are obtained, which must undergo a preliminary selection before the most intensive evaluation to identify the lines to be recombined. [ABSTRACT FROM AUTHOR]

Proline plays adaptive roles in plant tolerance to cadmium (Cd)-induced stress, but many gaps remain to be elucidated as the responses triggered by exogenously supplied proline or endogenously overproduction are not well known. Thus, we assayed the nutritional status, metabolite profiling, and antioxidative responses in wild type and transgenic tobacco (Nicotiana tabacum L.) containing the P5CSF129A gene under control of the cauliflower mosaic virus (CaMV35S) or stress inducible rd29A promoters. The plants were exposed or unexposed to Cd (0 and 50 μmol L−1 CdCl2·H2O) for 24 and 72 h. The wild type plants were also treated with or without exogenous proline (1 mmol L−1). Plants supplied with exogenous proline exhibited lower Cd translocation from roots to leaves than plants overproducing proline, avoiding oxidative damages in the leaves of these plants. Meanwhile, tobacco overproducing proline was less susceptible to Cd-induced nutritional changes than wild type plants and presented better metabolic adjustment under Cd exposure compared to plants supplied with exogenous proline. Plants overproducing proline increased the synthesis of sugars and organic acids under Cd exposure, which contributed to absence of oxidative stress, since both superoxide dismutase and catalase were not active against Cd-induced oxidative stress in these genotypes. Plants overproducing proline under the control of rd29A presented higher proline concentration in comparison to the CaMV35S promoter. With exception of rd29A plants that presented high proline and reduced glutathione (GSH) concentrations, the other plants presented an inverse correlation between proline and GSH synthesis after 72 h of Cd exposure. [ABSTRACT FROM AUTHOR]

The antimicrobial properties of silver and enhanced reactivity when applied in a nanoparticle form (AgNPs) led to their growing utilization in industry and various consumer products, which raises concerns about their environmental impact. Since AgNPs are prone to transformation, surface coatings are added to enhance their stability. AgNP phytotoxicity has been mainly attributed to the excess generation of reactive oxygen species (ROS), leading to the induction of oxidative stress. Herein, in vitro-grown tobacco (Nicotiana tabacum) plants were exposed to AgNPs stabilized with either polyvinylpyrrolidone (PVP) or cetyltrimethylammonium bromide (CTAB) as well as to ionic silver (AgNO3), applied in the same concentrations, either alone or in combination with cysteine, a strong silver ligand. The results show a higher accumulation of Ag in roots and leaves after exposure to AgNPs compared to AgNO3. This was correlated with a predominantly higher impact of nanoparticle than ionic silver form on parameters of oxidative stress, although no severe damage to important biomolecules was observed. Nevertheless, all types of treatments caused mobilization of antioxidant machinery, especially in leaves, although surface coatings modulated the activation of its specific components. Most effects induced by AgNPs or AgNO3 were alleviated with addition of cysteine. [ABSTRACT FROM AUTHOR]