Volatile substances for inhibiting growth of fusarium oxysporum by increasing H3K14ac levels

Abstract

The invention belongs to the technical field of using L-menthol as a novel plant source HDACi for inhibiting fungal growth, discloses a volatile substance for inhibiting the growth of fusarium oxysporum by improving the H3K14ac level and a method thereof, and finds that the monomer component L-menthol of mint volatile oil can inhibit the growth of fusarium oxysporum as a volatile small molecule; it is found that the histone deacetylation level of fusarium oxysporum is improved by the mint volatile oil monomer component L-menthol; the monomer component L-menthol of the mint volatile oil can inhibit the activity of histone deacetylase of fusarium oxysporum and inhibit the expression of key genes in a glycolysis / gluconeogenesis pathway.

Description

Technical Field

[0001] The invention belongs to the technical field of novel plant-derived HDACi using L-menthol as a fungal growth inhibitor, and in particular relates to a volatile substance and a method for inhibiting the growth of Fusarium oxysporum by increasing the H3K14ac level. Background Art

[0002] As the key pathogenic factor of Panax notoginseng root rot, Fusarium oxysporum invades the roots through soil media during the cultivation of Panax notoginseng, causing root tissue ulceration, posing a serious threat to the growth and development and final quality of Panax notoginseng. In view of this, Panax notoginseng growers widely use chemical fungicides such as methyl thiophanate, carbendazim and genkang to effectively curb the spread of pathogens and manage diseases. However, long-term reliance on such chemical agents not only causes pathogens to develop drug resistance, but also causes environmental pollution problems and may pose a potential threat to non-target biological populations. In this context, the research and development and application of green pesticides have become an important trend to promote the sustainable development of modern agriculture in my country and even the world. At present, the innovation of green pesticides focuses on the identification of new targets and the continuous exploration of chemical entities, aiming to develop new varieties of pesticides that are target-oriented, highly efficient, low-toxic and low-environmental risk. Significant progress has been made in this field, and pesticide technology and products are undergoing profound changes. Despite this, my country still faces the challenge of scarcity of original targets, which limits the independent research and development capabilities of efficient and low-risk small molecule pesticides based on molecular design technology. Therefore, strengthening the discovery and utilization of original targets and combining them with advanced molecular design strategies has become an urgent need to promote green pesticide innovation and promote green agricultural development.

[0003] Histone deacetylation is one of the most critical epigenetic mechanisms in eukaryotes, which can regulate various genes involved in fungal growth, development and response to environmental stress. HDAC inhibitors (HDACi) are usually used to study the molecular mechanism of histone deacetylation or promote the treatment of human diseases by hindering the catalytic activity of histone deacetylase. Inhibition of histone deacetylase has been shown to have a good therapeutic effect on many cancers through a large number of studies. In recent years, there are relatively few types of HDAC inhibitors currently on the market, which are mainly related to the field of medical technology and less used in the prevention and control of agricultural pests and diseases. Volatile substances play an important role in the biological control of agricultural diseases. However, there is no research on how volatile small molecules regulate histone deacetylation as HDACi and how they affect fungal growth and development, which has high research value and research space.

[0004] Through the above analysis, the problems and defects of the prior art are as follows:

[0005] There is no research on how volatile small molecules regulate histone deacetylation in Fusarium oxysporum and affect the growth and development of the fungus. Summary of the invention

[0006] In view of the problems existing in the prior art, the present invention provides a volatile substance for inhibiting the growth of Fusarium oxysporum by increasing the level of H3K14ac.

[0007] The present invention is achieved by a method for inhibiting the growth of Fusarium oxysporum by increasing the level of H3K14ac, comprising:

[0008] Step 1, sample processing:

[0009] 2 g of Fusarium oxysporum hyphae were added to a conical flask containing 40 mL of L-menthol solution of different concentrations, and the conical flask was placed in a constant temperature incubator at 28°C for constant temperature shaking culture for 24 h, and then the sample was collected and ground into powder using a cryogenic grinder for later use;

[0010] Step 2: Prepare histones of the treated samples according to the instructions of the histone extraction kit.

[0011] Furthermore, the L-menthol is 1 mmol / L and 2 mmol / L.

[0012] Furthermore, the histones of the treated sample are prepared according to the instructions of the histone extraction kit:

[0013] 15% sodium dodecyl sulfate polyacrylamide gel (SDSPAGE) was used for separation, and antiH3K14ac, antiH3K27ac, antiH3K9ac and antiH3 antibodies were used for immunoblotting to detect H3 lysine acetylation / status; AntiH3 immunoblot was used as a control; Image J software was used to quantify the band intensity.

[0014] Furthermore, the antiH3K14ac is: Jingjie, PTM157.

[0015] Furthermore, the antiH3K27ac is: Abcan, Cat#: ab24404.

[0016] Furthermore, the antiH3K9ac is: Abcam, Cat#: ab16635.

[0017] Furthermore, the antiH3 is: Jingjie, PTM1002.

[0018] Another object of the present invention is to provide a method for applying the above-mentioned volatile substances that inhibit the growth of Fusarium oxysporum by increasing the level of H3K14ac, comprising applying the treated Fusarium oxysporum mycelium sample to the agricultural field, inhibiting the growth of Fusarium oxysporum by increasing the level of H3K14ac, thereby reducing crop diseases.

[0019] Another object of the present invention is to provide a method for using the above-mentioned volatile substance for inhibiting the growth of Fusarium oxysporum by increasing the level of H3K14ac, comprising using the volatile substance as a food additive or preservative to inhibit the growth of Fusarium oxysporum in food by increasing the level of H3K14ac, thereby extending the shelf life of the food.

[0020] Another object of the present invention is to provide a method for using the above-mentioned volatile substance for inhibiting the growth of Fusarium oxysporum by increasing the level of H3K14ac, including applying the volatile substance to the field of biomedicine to inhibit the growth of Fusarium oxysporum by increasing the level of H3K14ac, thereby developing new antifungal drugs or treatment methods.

[0021] In combination with the above technical solutions and the technical problems solved, the advantages and positive effects of the technical solutions to be protected by the present invention are as follows:

[0022] First, the present invention finds that L-menthol, a monomer component of peppermint volatile oil, as a volatile small molecule, can inhibit the growth of Fusarium oxysporum; finds that L-menthol, a monomer component of peppermint volatile oil, increases the histone deacetylation level of Fusarium oxysporum; L-menthol, a monomer component of peppermint volatile oil, can inhibit the histone deacetylase activity of Fusarium oxysporum, thereby inhibiting the expression of key genes in the glycolysis / gluconeogenesis pathway.

[0023] Second, the expected benefits and commercial value of the technical solution of the present invention after transformation are:

[0024] Reduce the use of pesticides and improve crop yield and quality: Traditional pesticides have residues and environmental pollution problems when preventing and controlling crop diseases. L-menthol, as a natural plant-derived HDACi, kills pathogens in a targeted manner with high efficiency. This feature can not only effectively reduce crop diseases, but is also expected to significantly reduce the use of traditional pesticides, thereby increasing crop yield and quality and increasing farmers' income. At the same time, high-quality agricultural products have greater market demand, bringing higher sales prices, thereby increasing farmers' income.

[0025] Promoting sustainable agricultural development: Promoting the use of L-menthol as a new natural plant-based pesticide will help promote the development of agriculture in a green, ecological and sustainable direction, which is in line with the trends and requirements of global agricultural development.

[0026] Broad market demand: As people's awareness of food safety and environmental protection increases, the demand for natural, non-toxic and environmentally friendly pesticides is growing. As a new plant-based HDACi, L-menthol has the advantages of low toxicity, environmental protection and easy degradation. It can effectively reduce the risk of crop and environmental pollution and has broad market prospects in the field of green agriculture.

[0027] High technical barriers: At present, there are relatively few studies on the application of L-menthol in the agricultural field, and the technical barriers are relatively high. Once the technology is successfully transformed and applied to agricultural production, it will form a strong technical advantage and competitive advantage.

[0028] Industrial chain extension: The industrial production of L-menthol can be combined with agriculture, medicine, daily chemicals and other fields to form a complete industrial chain. Through technology transformation and industrial upgrading, it can drive the development of related industries and improve economic benefits.

[0029] Third, the technical solution of the present invention fills the technical gap in the industry at home and abroad:

[0030] Technical status at home and abroad: There is little information directly mentioning L-menthol as an invention of HDACi pesticide technology, but considering the wide range of effects of HDACi in organisms, including regulating gene expression and cell proliferation, L-menthol, as a natural active substance, may affect the growth and development of fungi by inhibiting the activity of HDACs (histone deacetylases), thereby showing the potential to inhibit fungal growth. In the field of pesticides, finding new pesticides that are highly efficient, low-toxic and environmentally friendly has always been a hot topic of research. Although traditional chemical pesticides are effective, they often have problems such as environmental pollution and ecological damage. Therefore, the development of new plant-based pesticides is of great significance.

[0031] Filling the technological gap: If the invention of L-menthol as a new plant-based HDACi pesticide technology for inhibiting fungal growth has been realized and successfully applied in actual production, then this invention will undoubtedly fill the technological gap in this field at home and abroad. It not only provides a new option for the pesticide industry, but also provides strong support for the sustainable development of agriculture.

[0032] Fourth, the technical solution of the present invention solves the technical problems that people have been eager to solve but have never been successful:

[0033] In the Chinese medicinal materials planting industry, some Chinese medicinal materials such as Paris polyphylla and Panax notoginseng are very susceptible to root rot pathogens such as Fusarium oxysporum, resulting in a serious decline in yield and quality. If it can be applied to Chinese medicinal materials planting, the occurrence of root rot can be prevented and controlled, and the quality and yield of Chinese medicinal materials can be improved, which will promote the innovative development of the Chinese medicinal materials industry. In crop planting, the use of chemical pesticides can be reduced or avoided, pesticide residues can be reduced, and crop quality can be improved.

[0034] Fifth, the technical solution of the present invention overcomes technical prejudice:

[0035] As a new type of enzyme inhibitor, HDACi is still in the exploratory stage in the application of pesticides, and its application scope and effect may be subject to certain restrictions. With the continuous deepening of research on HDACi pesticides, people have a more comprehensive understanding of its application potential and advantages in the field of pesticides. As one of the representatives of HDACi pesticides, L-menthol has been verified to inhibit fungal growth and has shown good application prospects in practical applications. This will help break the restrictions on the application of HDACi pesticides and promote their widespread application in the field of pesticides. BRIEF DESCRIPTION OF THE DRAWINGS

[0036] Figure 1 It is a flow chart of a method for producing volatile substances that inhibit the growth of Fusarium oxysporum by increasing the level of H3K14ac provided in an embodiment of the present invention.

[0037] Figure 2 This is a graph showing the effect of increasing different concentrations of L-menthol on the hyphae growth of Fusarium oxysporum provided by an embodiment of the present invention.

[0038] Figure 3 This is a diagram showing the effect of increasing different concentrations of L-menthol on H3K14ac provided by an embodiment of the present invention.

[0039] Figure 4 3 is a graph showing the effect of increasing L-menthol on FoRPD3 enzyme activity provided by an embodiment of the present invention.

[0040] Figure 5 This is a diagram showing the molecular docking results of improving L-menthol and FoRPD3 provided by an embodiment of the present invention.

[0041] Figure 6 This is a verification diagram of RTqPCR and ChIPqPCR in the L-menthol treatment group provided by an embodiment of the present invention. DETAILED DESCRIPTION

[0042] In order to make the purpose, technical solution and advantages of the present invention more clearly understood, the present invention is further described in detail below in conjunction with the embodiments. It should be understood that the specific embodiments described herein are only used to explain the present invention and are not used to limit the present invention.

[0043] like Figure 1 As shown, a method for producing volatile substances that inhibit the growth of Fusarium oxysporum by increasing the level of H3K14ac provided by an embodiment of the present invention comprises the following steps:

[0044] S101, sample processing:

[0045] 2 g of Fusarium oxysporum hyphae were added to a conical flask containing 40 mL of L-menthol solution of different concentrations, and the conical flask was placed in a constant temperature incubator at 28°C for constant temperature shaking culture for 24 h, and then the sample was collected and ground into powder using a cryogenic grinder for later use;

[0046] S102, preparing histones of the treated sample according to the instructions of the histone extraction kit.

[0047] The L-menthol provided in the embodiment of the present invention is 1 mmol / L and 2 mmol / L.

[0048] The histones of the sample after treatment are prepared according to the instructions of the histone extraction kit provided in the embodiment of the present invention:

[0049] 15% sodium dodecyl sulfate polyacrylamide gel (SDSPAGE) was used for separation, and antiH3K14ac, antiH3K27ac, antiH3K9ac and antiH3 antibodies were used for immunoblotting to detect H3 lysine acetylation / status; AntiH3 immunoblot was used as a control; Image J software was used to quantify the band intensity.

[0050] The antiH3K14ac provided in the embodiment of the present invention is: Jingjie, Cat#: PTM157.

[0051] The antiH3K27ac provided in the embodiment of the present invention is: Abcan, Cat#: ab24404.

[0052] The antiH3K9ac provided in the embodiment of the present invention is: Abcam, Cat#: ab16635.

[0053] The antiH3 provided in the embodiment of the present invention is: Jingjie, Cat#: PTM1002.

[0055] Example 1: L-Menthol Treatment of Fusarium oxysporum

[0056] L-menthol was diluted by two-fold dilution method and added to 20 mL of potato dextrose agar (PDA) medium (final monomer concentration gradient was 8, 4, 2, 1, 0.5, 0.25 mmol / L). The colony diameter was measured once every 24 h by the "cross" method until the 7th day, and the obtained values ​​were plotted into a line graph to calculate the half inhibition concentration (EC50).

[0057] The minimum inhibitory concentration (MIC) value was determined by 96-well plate method and spore suspension was prepared. The spore concentration was adjusted to 2×10 5The initial concentration of L-menthol was 4 mg / mL, and L-menthol was diluted into eight concentration gradients by two-fold dilution. The 96-well plate was incubated at 28°C for 20 h, and the absorbance of each well was measured at a wavelength of 595 nm to calculate the MIC value of L-menthol.

[0058] Example 2: Effect of L-menthol on histone H3K14ac of Fusarium oxysporum

[0059] Sample treatment: Take 2 g of Fusarium oxysporum mycelium and add it into a conical flask containing 40 mL of L-menthol solution (1 mmol / L and 2 mmol / L) of different concentrations. Place the conical flask in a constant temperature incubator at 28°C for constant temperature shaking incubation for 24 hours, then collect the sample and grind it into powder using a low-temperature grinder for later use.

[0060] Histone extraction kit The histones of the treated samples were prepared according to the instructions. 15% SDS PAGE gel was used for separation, and antiH3K14ac (Jingjie, PTM157), antiH3K27ac (Abcan, Cat#: ab24404), antiH3K9ac (Abcam, Cat#: ab16635) and antiH3 (Jingjie, PTM1002) antibodies were used for immunoblotting to detect H3 lysine acetylation / status. AntiH3 immunoblotting was used as a control. The band intensity was quantified using Image J software.

[0061] Example 3: Effect of L-menthol on FoRPD3 enzyme activity

[0062] In order to determine the activity of FoRPD3, we used gene recombination technology to transfer the target gene into an expression vector (pET28a, kan+), and expressed the protein through Escherichia coli (BL21). The concentration of the expressed protein was determined using the BCA method (BCA protein concentration determination kit SK1070). Fluorimetric HDAC Activity AssayKit*Green Fluorescence* measures the activity of histone HDAC deacetylase.

[0063] Example 4: Molecular docking of L-menthol and FoRPD3

[0064] Autodock Vina was used for molecular docking simulation. The 3D structure of FoRPD3 protein was downloaded from the Unipot official website (https: / / www.uniprot.org / uniprotkb / Q9FE17 / entry#names_and_taxonomy), and the 3D structure of L-menthol was downloaded from the Pubchem (https: / / pubchem.ncbi.nlm.nih.gov / ) database. The protein receptor was dehydrated, hydrogenated, and energy minimized using the AutoDock Tool software. Molecular docking was performed and the results were calculated using Autodock Vina, and the results were visualized using PyMOL and Ligplot software.

[0065] Example 5: RTqPCR and ChIPqPCR analysis of Fusarium oxysporum treated with L-menthol

[0066] Sample treatment: Take 2 g of Fusarium oxysporum mycelium and add it into a conical flask containing 40 mL of L-menthol solution (1 mmol / L and 2 mmol / L) of different concentrations. Place the conical flask in a constant temperature incubator at 28°C for constant temperature shaking incubation for 24 hours, then collect the sample and grind it into powder using a low-temperature grinder for later use.

[0067] Take 2g of Fusarium oxysporum powder treated with monomers of different concentrations, extract total RNA of the sample with Trizol reagent, divide the above fungal sample powder into 1.5mL EP tubes, and immediately add Trizol reagent to fully lyse. Then add chloroform, isopropanol and ethanol to remove proteins and DNA. Finally, elute with DEPC water. Use a reverse transcription kit to synthesize the first-strand cDNA from total RNA. QTUB is used as a reference gene, and RTqPCR is performed in the CFXManager 3.1 system using the SYBR Premix Ex Taq kit (TaKaRa Biomedicals). Use 2 △△Cq Methods The relative expression of genes was analyzed.

[0068] Take 2g of Fusarium oxysporum powder after monomer treatment with different concentrations, add 30mL 1% formaldehyde solution and shake for cross-linking for 5min, then add 2.5M glycine and shake for 5min to terminate cross-linking. Add 30mL extraction buffer I and mix well, then centrifuge at low temperature and discard the supernatant, repeat the washing twice, then use 300μL extraction buffer III for density gradient centrifugation, discard the supernatant, and obtain the cell nucleus. Resuspend the cell nucleus with cell nucleus lysis buffer and perform high-power ultrasound at 4℃ for 50min (30s on, 30s off). Take 10μL of the ultrasound product to de-crosslink and detect whether the DNA fragments after ultrasound are distributed around 500bp. If the DNA detection quality meets the standard, the ChIP experiment can be carried out. The ultrasonic sample was added to the Protein A magnetic beads (with bound antibody H3K14ac) and incubated overnight in a 4°C refrigerator. The complex magnetic beads were washed with low salt, high salt, LiCl and TE buffer in sequence. Finally, the binding product was eluted with 250 μL elution buffer in a constant temperature metal bath at 65°C for 25 min (mixed every 2 min). The supernatant was aspirated into a clean EP tube, and the process was repeated twice for a total of 500 μL. 20 μL 5M NaCl was added to the eluted collection solution and cross-linked overnight in a metal bath at 65°C. Finally, the cross-linked product was removed and purified to obtain DNA. ChIPqPCR was performed in the CFXManager 3.1 system (BioRad) using the SYBR Premix Ex Taq kit (TaKaRa Biomedicals). The relative expression of genes was analyzed using the Percent Input method.

[0069] Figure 2 The results show that L-menthol has a good inhibitory effect on Fusarium oxysporum, with an EC50 of 2.489mmol / L and a commercial fungicide, oxadiazine, with an EC50 of 0.428mmol / L. It has similar inhibitory effects at higher concentrations, and is even better than oxadiazine. The minimum inhibitory concentration of L-menthol on Fusarium oxysporum is 1.95mmol / L.

[0070] Figure 3 The figure shows the effect of L-menthol on H3K14ac of Fusarium oxysporum. The study used western blot to detect the changes in H3K9ac, H3K14ac and H3K27ac levels in samples treated with L-menthol. As shown in the figure, after treatment with different concentrations of L-menthol, the abundance of H3K14ac increased, with significant differences, while H3K9ac and H3K27ac did not change significantly. The results showed that with the increase in the concentration of L-menthol, the abundance of H3K14ac in Fusarium oxysporum gradually increased, with significant differences, indicating that L-menthol can act as an external factor to affect histone H3K14ac modification.

[0071] Figure 4 The figure shows the effect of L-menthol on the enzyme activity of FoRPD3. First, the target protein FoRPD was expressed by prokaryotic protein expression. Compared with CK, the activity of FoRPD3 was significantly reduced after L-menthol treatment, which was 9344.4RFU / min / g. The activity of the commercial inhibitor (guqustatin A) on FoRPD3 was 9665.5RFU / min / g. The inhibitory effect of L-menthol on the enzyme activity of FoRPD3 was similar to that of the commercial inhibitor. L-menthol may affect the growth of Fusarium oxysporum by inhibiting the activity of FoRPD3 and regulating the modification level of H3K14ac.

[0072] Figure 5 This is the molecular docking diagram of L-menthol and FoRPD3. PyMOL was used to visualize some of the docking results. The binding energy of the binding complex of FoRPD3 protein and L-menthol is 5.3 kcal / mol. L-menthol is wrapped in the active pocket of FoRPD3 protein, and has hydrophobic interactions with amino acid residues such as Leu282, Gly160, and His189, which promotes the formation of the binding and improves the stability of the binding.

[0073] Figure 6 The RTqPCR and ChIPqPCR validation diagrams of gene expression in the L-menthol treatment group. ChIPqPCR further verified that after L-menthol treatment, the H3K14ac enrichment multiples of the key genes FOBCDRAFT_215442, FOBCDRAFT_36895, FOBCDRAFT_173064, and FOBCDRAFT_51813 in the glycolysis / gluconeogenesis pathway of Fusarium oxysporum significantly increased, which was consistent with the results of protein immunoblotting (WB) detection, which was an increase. The RTqPCR experiment showed that L-menthol can significantly inhibit the expression of these genes, thereby affecting the function of glycolysis / gluconeogenesis. It further shows that L-menthol can inhibit the activity of the histone deacetylase FoRPD3 in Fusarium oxysporum, thereby increasing the modification level of H3K14ac, and finally leading to the inhibition of gene expression in the glycolysis / gluconeogenesis pathway of Fusarium oxysporum.

[0074] Application areas:

[0075] Control of fungal diseases of crops: L-menthol has a strong bactericidal effect and can prevent and control fungal diseases on crops, such as root rot, leaf spot, powdery mildew, etc., by inhibiting the growth of fungi.

[0076] Crop pest control: In addition to its fungicidal effect, L-menthol can also be used to control agricultural pests such as aphids and caterpillars.

[0077] Forestry and horticultural disease control: L-Menthol can also be used to control diseases and insect pests on trees and flowers, protecting the healthy growth of plants.

[0078] Eco-friendly pesticides: Compared with traditional chemical pesticides, L-menthol, as a plant-based pesticide, does not pollute the ecological environment and meets the requirements of green agriculture and sustainable development.

[0079] Related Products:

[0080] L-Menthol pesticide preparation: L-Menthol is mixed with other auxiliary ingredients (such as surfactants, stabilizers, etc.) to make an easy-to-spray pesticide preparation for controlling diseases and insect pests on crops.

[0081] Compound pesticides: L-menthol is compounded with other active ingredients of pesticides (such as other botanical pesticides, biological pesticides or low-toxic chemical pesticides) to form compound pesticide products with a broader spectrum of fungicides and insecticides.

[0082] Plant growth regulators: Utilize the regulatory effect of L-menthol on plant growth to develop plant growth regulators that can promote crop growth and improve stress resistance.

[0083] Through antibacterial tests, L-menthol has a good inhibitory effect on Fusarium oxysporum, and is stronger than the commercial fungicide oxazolidinone at a higher concentration (8mmol / L). Further verification of its mechanism of action showed that L-menthol has a significant inhibitory effect on the activity of FoRPD3 of Fusarium oxysporum, and the effect is equivalent to that of the commercial inhibitor (guqustatin A), thereby causing high acetylation of H3K14ac in Fusarium oxysporum. Molecular docking simulation verification showed that L-menthol was wrapped in the active pocket of FoRPD3 protein, and there was a hydrophobic interaction with amino acid residues such as Leu282, Gly160, and His189, which was the main binding site with a binding energy of 5.3kcal / mol and good stability. RTqPCR and ChIPqPCR results further proved that after L-menthol treatment, the genes of the glycolysis / gluconeogenesis pathway of Fusarium oxysporum were at a high H3K14ac acetylation level, and gene expression was inhibited. In summary, L-menthol can inhibit the activity of histone deacetylase in pathogenic fungi, leading to hyperacetylation of histones and inhibiting gene expression.

[0084] L-Menthol is derived from natural plants. Compared with chemically synthesized pesticides, it has lower toxicity and is environmentally friendly. This provides an important safety guarantee for it as a new plant-based pesticide. In addition, L-menthol is easily degraded and will not remain in the environment for a long time, reducing potential harm to the ecosystem. L-Menthol can be efficiently prepared by chemical synthesis or biocatalysis, and has the advantages of mild reaction conditions, high selectivity, and high product purity. It is green, environmentally friendly, and efficient in agricultural production, and has great potential as a new plant-based HDACi pesticide to inhibit fungal growth.

[0085] The above description is only a specific implementation mode of the present invention, but the protection scope of the present invention is not limited thereto. Any modification, equivalent substitution and improvement made by any technician familiar with the technical field within the technical scope disclosed by the present invention and within the spirit and principle of the present invention should be covered by the protection scope of the present invention.

Claims

1. A volatile substance for inhibiting the growth of Fusarium oxysporum, characterized in that: This substance achieves its inhibitory effect by increasing the level of H3K14ac. Its main component is L-menthol, which is in the form of colorless needle-shaped crystals or granules and has a refreshing mint aroma. The concentration of L-menthol has a significant effect on the increase in the level of H3K14ac.

2. The volatile substance according to claim 1, characterized in that The concentration of L-menthol is 1 mmol / L and 2 mmol / L, which is used to effectively increase the H3K14ac level in Fusarium oxysporum.

3. The volatile substance according to claim 1, characterized in that The main component of the L-menthol is cyclohexane, which contains three asymmetric carbon atoms and has multiple cis and trans isomers. Each isomer has different optical isomers, including L-isomer, D-isomer and DL-isomer. In nature, it mainly exists in the essential oil of mint plants in the form of L-isomer.

4. The volatile substance according to claim 1, characterized in that The chemical property of the L-menthol is relatively stable, and it can volatilize together with steam and participate in a variety of chemical reactions, including hydrogenation reaction and esterification reaction. This chemical property makes it have high stability and effectiveness in application.

5. A method for inhibiting the growth of Fusarium oxysporum by increasing the level of H3K14ac, characterized in that: The following steps are involved: Take 2 g of Fusarium oxysporum mycelium and add it into a conical flask containing 40 mL of L-menthol solution of different concentrations; The conical flask was placed in a constant temperature incubator at 28°C with constant temperature shaking for 24 hours; Collect the samples and grind them into powder using a cryogenic grinder for later use; Prepare histones from the treated samples according to the instructions of the histone extraction kit.

6. The method according to claim 5, characterized in that The concentration of L-menthol is 1 mmol / L and 2 mmol / L, which is used to effectively increase the H3K14ac level in Fusarium oxysporum.

7. The method according to claim 5, characterized in that The extracted histones were separated by 15% SDS PAGE gel, and the H3 lysine acetylation status was detected by immunoblotting using antiH3K14ac, antiH3K27ac, antiH3K9ac and antiH3 antibodies. AntiH3 immunoblotting was used as a control, and the band intensity was quantitatively analyzed using Image J software.

8. The method according to claim 5, characterized in that By using L-menthol as the main ingredient, H3K14ac levels were significantly increased, thereby effectively inhibiting the growth of Fusarium oxysporum, wherein the L-menthol acts as a FoArginase_HDAC activator.

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