Development of a nornicotine-reduced flue-cured tobacco line via EMS mutagenesis of nicotine N-demethylase genes.
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ABSTRACT:
SUBMITTER: Song Z
PROVIDER: S-EPMC7053972 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:Up to now, the mechanism of the effect of topping on tobacco hormone regulation is not clear, and most studies on plant hormone signal transduction pathways rely on gene or transcriptional pathways. In this study, the regulatory mechanism of hormones in roots and leaves of topped and untopped tobacco was studied at the protein level.
2023-10-19 | PXD043495 | Pride
Project description:Nicotine, the most abundant pyridine alkaloid in cultivated tobacco (Nicotiana tabacum L.), is a potent inhibitor of insect and animal herbivory and a neurostimulator of human brain function. Nicotine biosynthesis is controlled developmentally and can be induced by abiotic and biotic stressors via a jasmonic acid (JA)-mediated signal transduction mechanism involving members of the APETALA 2/ethylene-responsive factor (AP2/ERF) and basic helix-loop-helix (bHLH) transcription factor (TF) families. AP2/ERF and bHLH TFs work combinatorically to control nicotine biosynthesis and its subsequent accumulation in tobacco leaves. Here, we demonstrate that overexpression of the tobacco NtERF32, NtERF221/ORC1, and NtMYC2a TFs leads to significant increases in nicotine accumulation in T2 transgenic K326 tobacco plants before topping. Up to 9-fold higher nicotine production was achieved in transgenics overexpressing NtERF221/ORC1 under the control of a constitutive GmUBI3 gene promoter compared to wild-type plants. The constitutive 2XCaMV35S promoter and a novel JA-inducible 4XGAG promoter were less effective in driving high-level nicotine formation. Methyljasmonic acid (MeJA) treatment further elevated nicotine production in all transgenic lines. Our results show that targeted manipulation of NtERF221/ORC1 is an effective strategy for elevating leaf nicotine levels in commercial tobacco for use in the preparation of reduced risk tobacco products for smoking replacement therapeutics.
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Project description:To study the relationship between the diversity of the surface microbial community and tobacco flavor, and to improve tobacco quality using microorganisms. The microbial community composition and diversity of 14 samples of flue-cured tobacco from tobacco-producing areas in Yunnan with varying grades were analyzed by high-throughput sequencing. PICRUSt was used for predicting microbial functions. A strain of Bacillus amyloliquefaciens W6-2 with the ability to degrade pectin was screened from the surface of flued-cured tobacco leaves from Yunnan reroasted tobacco leave. The enzyme preparation was prepared through fermentation and then applied for treating flue-cured tobacco. The improvement effect was evaluated by measuring the content of macromolecule and the changes in volatile components, combined with sensory evaluations. The bacterial communities on the surface of flue-cured tobacco exhibited functional diversity, consisting primarily of Variovorax, Pseudomonas, Sphingomonas, Burkholderia, and Bacillus. These bacterial strains played a role in the aging process of flue-cured tobacco leaves by participating in amino acid metabolism and carbohydrate metabolism. These metabolic activity converted complex macromolecules into smaller molecular compounds, ultimately influence the smoking quality and burning characteristics of flue-cured tobacco. The pectinase preparation produced through fermentation using W6-2 has been found to enhance the aroma and sweetness of flue-cured tobacco, leading to improved aroma, reduced impurities, and enhanced smoothness. Additionally, the levels of pectin, cellulose, and hemicellulose decreased, while the levels of water-soluble sugar and reducing sugar increased, and the contents of esters, ketones, and aldehydes increased, and the contents of benzoic acid decreased. The study revealed the correlation between surface microorganisms and volatile components of Yunnan tobacco leaves, and the enzyme produced by the pectin-degrading bacteria W6-2 effectively improved the quality of flue-cured tobacco.
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Project description:Fungal communities are considered to be critically important for crop health and soil fertility. However, our knowledge of the response of fungal community structure to the continuous cropping of flue-cured tobacco is limited, and the interaction of soil fungal communities under different cropping systems remains unclear. In this study, we comparatively investigated the fungal abundance, diversity, and community composition in the soils in which continuous cropping of flue-cured tobacco for 3 years (3ys), 5 years (5ys), and cropping for 1 year (CK) using quantitative polymerase chain reaction and high-throughput sequencing technology. The results revealed that continuous cropping of flue-cured tobacco changed the abundance of soil fungi, and caused a significant variation in fungal diversity. In particular, continuous cropping increased the relative abundance of Mortierellales, which can dissolve mineral phosphorus in soil. Unfortunately, continuous cropping also increased the risk of potential pathogens. Moreover, long-term continuous cropping had more complex and stabilize network. This study also indicated that available potassium and available phosphorous were the primary soil factors shifting the fungal community structure. These results suggested that several soil variables may affect fungal community structure. The continuous cropping of flue-cured tobacco significantly increased the abundance and diversity of soil fungal communities.
| S-EPMC7669846 | biostudies-literature
Project description:A range of endemic Nicotiana species are chewed as a smokeless tobacco by several Aboriginal populations of Australia. In tobacco research, nicotine to nornicotine conversion is important because nornicotine lowers tobacco quality and is detrimental to health. A diverse group of cytochrome P450 genes with different transcriptional regulations are involved in this conversion. The primary aims of this study were to quantify the pyridine alkaloids and investigate nicotine to nornicotine conversion in laboratory-grown Australian Nicotiana spp. Nicotine, nornicotine, anatabine, anabasine, myosmine and cotinine were quantified in fresh leaves of 24 out of the 26 recognised Australian Nicotiana taxa. Conserved regions of CYP82E related genes were PCR amplified in all studied taxa. The conversion process in fresh leaves was compared with that in leaves that underwent a simulated curing process for species that we identified as being high converters (N. cavicola, N. goodspeedii, N. velutina) and low converters (N. benthamiana, N. excelsior, N. gossei). Agarose gel electrophoretic analysis of CYP82E related genes obtained from the PCR amplification of the cDNA in fresh versus leaves with simulated curing showed about a 3-fold increase in transcript accumulation levels in cured leaves of the high converter species, while the transcript accumulation in N. gossei and N. excelsior maintained a steady basal level and increased by a small amount in N. benthamiana. This suggests the presence of functional loci that are triggered by curing in only high converter species and indicates a potential risk for chewers of high converter species.
| S-EPMC5727613 | biostudies-literature
Project description: Not available
| S-EPMC10568975 | biostudies-literature
Project description:Starch is an essential factor affecting the quality of flue-cured tobacco, and high starch content can affect the sensory quality and safety. Recently, the degradation of macromolecules in tobacco raw materials by using additional microorganisms to improve their intrinsic quality and safety has become a new research hotspot in the tobacco industry. However, the technical maturity and application scale are limited. Our study analyzed the correlation between microbial community composition and volatile components on the surface of tobacco leaves from 14 different grades in Fujian tobacco-producing areas. The PICRUSt software was utilized to predict the function of the microbial community present in tobacco leaves. Furthermore, dominant strains that produced amylase were screened out, and an enzyme solution was prepared to enhance the flue-cured tobacco flavor. Changes in the content of macromolecules and volatile components were determined, and sensory evaluations were conducted to assess the overall quality of the tobacco leaves. The results showed that the dominant bacterial genera on the surface of Fujian tobacco leaves were Variovorax, Sphingomonas, Bacillus, etc. Bacillus was positively correlated with various volatile components, which contributed to the sweet and aromatic flavors of Fujian flue-cured tobacco. The main genetic functions of Fujian flue-cured tobacco surface bacteria were carbohydrate metabolism and amino acid metabolism. After treating flue-cured tobacco with an enzyme preparation prepared by the fermentation of Paenibacillus amylolyticus A17 #, the content of starch, pectin, and cellulose in flue-cured tobacco decreased significantly compared with the control group. Meanwhile, the content of total soluble sugar and reducing sugar was significantly increased, and the volatile aroma components, such as 3-hydroxy--damascone, 2,3-dihydro-3,5-dihydroxy-6-methyl-4 H-Pyran-4-one, ethyl palmitate, ethyl linolenic acid, etc., were significantly increased. The aroma quality and quantity of flue-cured tobacco were enhanced, while impurities were reduced. The smoke characteristics were improved, with increased fineness, concentration, and moderate strength. The taste characteristics were also improved, with reduced irritation and a better aftertaste. In conclusion, Bacillus, as the dominant genus in the abundance of bacterial communities on tobacco surfaces in Fujian, had an essential impact on the flavor of tobacco leaves by participating in carbohydrate metabolism and finally forming the unique flavor style of flue-cured tobacco in Fujian tobacco-producing areas. Paenibacillus amylolyticus A17 #, a target strain with amylase-producing ability, was screened from the surface of Fujian flue-cured tobacco. The enzyme preparation, produced by the fermentation of Paenibacillus amylolyticus A17 #, was utilized to reduce the content of macromolecules, increase the content of water-soluble total sugar and reducing sugar, and produce a variety of crucial volatile aroma components, which had a significant improvement on the quality of tobacco leaves.
| S-EPMC10335769 | biostudies-literature
Project description:BackgroundThe aging process in the tobacco production, as in other food industries, is an important process for improving the quality of raw materials. In the spontaneous aging, the complex components in flue-cured tobacco (FT) improve flavor or reduce harmful compounds through chemical reactions, microbial metabolism, and enzymatic catalysis. Some believed that tobacco-microbe played a significant part in this process. However, little information is available on how microbes mediate chemical composition to improve the quality of FT, which will lay the foundation for the time-consuming spontaneous aging to seek ways to shorten the aging cycle.ResultsComparing aged and unaged FT, volatile and non-volatile differential compounds (DCs) were multi-dimensionally analyzed with the non-targeted metabolomes based on UPLC-QTOP-MS (the ultra-performance liquid chromatography quadrupole time-of-flight mass spectrometry), GC-MS (gas chromatography-mass spectrometer) assisted derivatization and HP-SPME-GC/MS (headspace solid-phase micro-extraction assisted GC-MS). Products associated with the degradation pathways of terpenoids or higher fatty acids were one of the most important factors in improving FT quality. With the microbiome, the diversity and functions of microbial flora were analyzed. The high relative abundance function categories were in coincidence with DCs-related metabolic pathways. According to the correlation analysis, Acinetobacter, Sphingomonas and Aspergillus were presumed to be the important contributor, in which Aspergillus was associated with the highest number of degradation products of terpenoids and higher fatty acids. At last, the screened Aspergillus nidulans strain F4 could promote the degradation of terpenoids and higher fatty acids to enhance tobacco flavor by secreting highly active lipoxygenase and peroxidase, which verified the effect of tobacco-microbes on FT quality.ConclusionsBy integrating the microbiome and metabolome, tobacco-microbe can mediate flavor-related substances to improve the quality of FT after aging, which provided a basis for identifying functional microorganisms for reforming the traditional spontaneous aging.
| S-EPMC9377114 | biostudies-literature
Project description:Microorganisms present on the surface of tobacco leaves play a significant role in shaping the composition of the tobacco microbial ecosystem, which undergoes continuous changes throughout the curing process. In the present study, a total of four distinct tobacco curing periods were selected for sampling, namely the fresh, yellowing, leaf-drying, and stem-drying stages. The bacterial 16S rRNA gene sequences of the collected samples were subsequently analyzed to identify operational taxonomic units (OTUs). The findings indicated that the complete dataset of leaf microbial samples was clustered, resulting in the identification of 1,783 operational taxonomic units (OTUs). Furthermore, the analysis of diversity revealed a pattern of initially increasing and subsequently decreasing community diversity. Redundancy Analysis (RDA) and weighted gene correlation networks for analysis (WGCNA) were employed in conjunction with environmental factors to assign OTUs to 22 modules for functional analysis. Additionally, a classification model utilizing the random forest algorithm was utilized to identify seven marker microorganisms (Escherichia coli, Faecalibacterium prausnitzii, Faecalibacterium, Escherichia-Shigella, Peptostreptococcaceae, Peptostreptococcales-Tissierellales, and Proteobacteria) that exhibited discriminative characteristics across different time periods. This study aimed to investigate the dynamic changes in the bacterial community throughout the curing process and their impact on the community's function. Additionally, certain bacteria were identified as potential markers for detecting changes in the curing stage. These findings offer a novel opportunity to accurately regulate the curing environment, thereby enhancing the overall quality of tobacco leaf curing.
| S-EPMC10713716 | biostudies-literature
Project description:BackgroundLeaves of tobacco (Nicotiana tabacum L.) are flue-cured to use as a key industrial supply in various parts of the world. The quality of tobacco leaves is dependent on chemical components and their proportions. Generally, the stem attached to tobacco leaf is detached before curing. However, the leaf stem remains green for an extended period of time (as compared to leaf) during flue-curing. Hence, it is expected to affect the quality of tobacco's final product.ResultsTo understand the impact of the green stem of leaf on the metabolome of flue-cured tobacco, we employed a broad targeted metabolomics approach. We selected two tobacco cultivars (Yun87 and K326) and cultivated them in five geographic locations in China. For flue-curing, leaves were harvested without a stem (L) or with an attached stem (SPL). After metabolome analysis, a total of 1027 metabolites were annotated in these samples. A variable number of metabolites were differentially accumulated between both types of leaves (depending on geographic location or cultivar) representing an influence of environment or genotype. Interestingly, only 68 metabolites were differentially accumulated between L and SPL samples irrespective of the cultivar or geographic location. These differentially accumulated metabolites belonged to major groups of primary and secondary metabolites. We have discussed the importance of identified metabolites in terms of carbon, nitrogen, and polyphenolic metabolism.ConclusionThe present research is the first comprehensive description of several metabolites in tobacco leaves related to the contribution of leaf stem. The current study opens novel prospects for investigating the potential of such metabolites in improving the quality of flue-cured tobacco.
| S-EPMC9926566 | biostudies-literature