A chitin-rich engineered bacterium and its construction method and application

Abstract

The present invention belongs to the field of genetic engineering technology and specifically relates to an engineered bacterium with a high chitin content, its construction method, and its application. This invention uses genetic engineering to overexpress enzymes involved in chitin synthesis in filamentous fungi to construct an engineered bacterium. The resulting engineered bacterium has a high chitin content and can be used to produce high-strength mycelium material through conventional solid-state fermentation. This invention provides a practical solution for the production of high-strength mycelium material with controllable product quality, and has excellent application value.

Description

Technical Field

[0001] The present invention belongs to the technical field of genetic engineering, and in particular relates to an engineering bacterium with a high chitin content, a construction method thereof and an application thereof. Background Art

[0002] Mycelium material is made from fungal hyphae through extraction, cleaning, and processing. Its unique three-dimensional structure and numerous excellent properties have led to its widespread application in environmentally friendly materials, construction, clothing, and packaging. Mycelium, the fundamental unit of mycelium material, is composed of its own components, which collectively give it unique physical and chemical properties.

[0003] The cell wall, the outer barrier of the mycelium, is primarily composed of polysaccharides such as chitin and glucan. These polysaccharides form a complex three-dimensional network, endowing the mycelium with a stable morphology and structure, enabling it to maintain its integrity under various environmental conditions. Chitin, a major component of the mycelial cell wall, plays a number of important roles in mycelial materials: 1. Providing structural support and mechanical strength: Chitin forms a robust network within the cell wall, imparting excellent mechanical strength and stability to the mycelium, enabling it to withstand certain physical pressures and mechanical loads. 2. Influencing material properties: Chitin not only influences the material's hardness and strength, but also its flexibility, water absorption, and thermal stability. For example, chitin's polysaccharide network helps improve the mycelium's water absorption and retention, making it more hydrophilic and biocompatible in certain applications. 3. Contributing to the material's biodegradability: Chitin's excellent biodegradability allows the mycelium to naturally degrade after use, minimizing environmental pollution. This degradability is in line with the concept of sustainable development, making mycelium materials an environmentally friendly alternative; 4. Affects the interaction with other materials. In practical applications, mycelium materials often need to be used in combination with other materials. The presence of chitin will affect the interaction between mycelium and other materials. For example, the surface properties and chemical composition of chitin will affect the adhesion and compatibility of mycelium with other materials, thereby affecting the performance and application effects of the composite materials; 5. Biological activity and application potential. Chitin has multiple biological activities, such as antibacterial, antioxidant and immunomodulatory. These characteristics make mycelium materials have broad application potential in many fields such as medical treatment, agricultural production increase, and environmental protection management. For example, chitin can be used to make medical materials such as surgical sutures, hemostatic sheets and diaphragms, and can also be used as a medical and health product with functions such as regulating the intestinal environment.

[0004] Currently, mycelial materials are often prepared by exogenously adding chitin synthesis precursors during culture to induce the mycelium to produce more chitin, thereby improving the strength of the mycelial material. However, this method often requires precise control of the culture medium composition to achieve the desired induction effect, which increases the complexity of the process. Furthermore, excessive addition of precursors can lead to uneven internal structure of the mycelium, which may in turn reduce the mechanical properties of the mycelial material. Summary of the Invention

[0005] To address the above issues, the present invention provides an engineered bacterium with high chitin content, a method for constructing the bacterium, and its application. The engineered bacterium provided by the present invention has a high chitin content in its cell wall, and can produce high-strength mycelium materials under conventional solid-state fermentation conditions, thus having good application prospects.

[0006] In order to achieve the above-mentioned object of the invention, the present invention adopts the following technical solutions:

[0007] The first aspect of the present invention provides an engineered bacterium with a high chitin content. The engineered bacterium is constructed by genetically modifying a filamentous fungus as a starting strain; the genetic modification includes overexpressing chitin synthase (CHS), glutamine-fructose-6-phosphate transaminase (GFAT), and uridine diphosphate-N-acetylglucosamine pyrophosphorylase (UAP).

[0008] The present invention expresses GFAT, UAP, and CHS enzymes in the fungus, so that more energy metabolism is directed to chitin synthesis. As a result, the constructed engineered bacteria can significantly increase the chitin content in the cell wall under conventional fermentation conditions without the need for exogenous addition of inducers such as chitin synthesis precursors, thereby improving the mechanical strength of the mycelium material obtained by fermentation, providing a new approach to obtaining high-strength mycelium materials.

[0009] Preferably, the filamentous fungi include Ganoderma lucidum, Pleurotus ostreatus, Cordyceps militaris, Versicolor versicolor, and Flammulina velutipes.

[0010] Preferably, the chitin synthase is chitin synthase CHS-5 or CHS-8.

[0011] Preferably, the uridine diphosphate-N-acetylglucosamine pyrophosphorylase is uridine diphosphate-N-acetylglucosamine pyrophosphorylase UAP1.

[0012] Preferably, the method for overexpressing the chitin synthase, glutamine-fructose-6-phosphate transaminase and uridine diphosphate-N-acetylglucosamine pyrophosphorylase is: increasing the copy number of the encoding gene of each enzyme.

[0013] The second aspect of the present invention provides a method for improving the strength of mycelium materials, which comprises increasing the chitin, β-glucan and protein capable of binding to cell wall components in the mycelium cell wall.

[0014] Preferably, the method is to overexpress chitin synthase, glutamine-fructose-6-phosphate transaminase and uridine diphosphate-N-acetylglucosamine pyrophosphorylase.

[0015] A third aspect of the present invention provides an engineered Ganoderma lucidum with a high chitin content. The engineered Ganoderma lucidum is constructed using Ganoderma lucidum as a starting strain through genetic modification; the genetic modification comprises overexpressing chitin synthase CHS-5 or CHS-8, glutamine-fructose-6-phosphate transaminase GFAT, and uridine diphosphate-N-acetylglucosamine pyrophosphorylase UAP1, wherein the amino acid sequence of the chitin synthase CHS-5 is shown in GenBank: AZQ26802.1, the amino acid sequence of the chitin synthase CHS-8 is shown in GenBank: AZQ26805.1, the amino acid sequence of the glutamine-fructose-6-phosphate transaminase GFAT is shown in SEQ ID NO.1, and the amino acid sequence of the uridine diphosphate-N-acetylglucosamine pyrophosphorylase UAP1 is shown in SEQ ID NO.2.

[0016] Preferably, the nucleotide sequence of the gene encoding the chitin synthase CHS-5 is shown in GenBank: MH463055.1, the nucleotide sequence of the gene encoding the chitin synthase CHS-8 is shown in GenBank: MH463058.1, the nucleotide sequence of the gene encoding the glutamine-fructose-6-phosphate transaminase GFAT is shown in SEQ ID NO.3, and the nucleotide sequence of the gene encoding the uridine diphosphate-N-acetylglucosamine pyrophosphorylase UAP1 is shown in SEQ ID NO.4.

[0017] Illustratively, the present invention also provides a method for constructing the above-mentioned high-chitin-containing Ganoderma lucidum engineered bacteria, which specifically comprises the following steps:

[0018] S1, construction of recombinant plasmid 1303-GLP-chitin for chitin synthase CHS-5 or CHS-8, glutamine-fructose-6-phosphate transaminase GFAT, and uridine diphosphate-N-acetylglucosamine pyrophosphorylase UAP1;

[0019] S2. Transforming the recombinant plasmid 1303-GLP-chitin into Ganoderma lucidum protoplasts, and screening the obtained positive transformants to obtain the Ganoderma lucidum engineered bacteria with high chitin content.

[0020] When screening the positive transformants obtained in S2, positive transformants with significantly high expression of chitin synthase CHS-5 or CHS-8, glutamine-fructose-6-phosphate transaminase GFAT, and uridine diphosphate-N-acetylglucosamine pyrophosphorylase UAP1 are selected to obtain the engineered Ganoderma lucidum strain with high chitin content. "Significantly high expression" means that the degree of increase in gene expression level is significantly different as commonly understood in the art.

[0021] The fourth aspect of the present invention provides an engineered bacterium GL-chintin-19 with high chitin content, which is classified as Ganoderma lucidum ( Ganoderma lingzh i), deposited on May 21, 2025, at the General Microbiology Center of China Culture Collection Administration, address: No. 3, Yard 1, Beichen West Road, Chaoyang District, Beijing, with the deposit number CGMCC No. 42012.

[0022] The engineered strain GL-chintin-19 was constructed using a commercial Ganoderma lucidum strain as a starting point by overexpressing the chitin synthase CHS-5, the glutamine-fructose-6-phosphate transaminase GFAT, and the uridine diphosphate-N-acetylglucosamine pyrophosphorylase UAP1. Compared to the chitin content of the starting strain, the chitin content of the engineered strain GL-chintin-19 increased by 548%. The mycelial fiber sheath produced by the engineered strain GL-chintin-19 was 149% stronger than that produced by the starting strain.

[0023] The fifth aspect of the present invention provides the use of the above-mentioned engineered bacteria, engineered Ganoderma lucidum bacteria, and engineered bacteria GL-chintin-19 in the production of mycelium materials.

[0024] A sixth aspect of the present invention provides a method for producing mycelium fiber skin, wherein the mycelium fiber skin is obtained by solid-state fermentation using the above-mentioned engineered bacteria or the engineered bacteria GL-chintin-19.

[0025] The method specifically includes the following operations: spraying the mycelial suspension of the above-mentioned engineered bacteria or the engineered bacteria GL-chintin-19 on the surface of the solid fermentation substrate, culturing at a constant temperature of 28-30°C and RH 60%-70%, and separating and removing the mycelial skin above the substrate after the cultivation cycle to obtain the mycelial fiber skin.

[0026] Preferably, the mycelium fiber skin is obtained by solid-state fermentation using the above-mentioned engineered bacteria GL-chintin-19.

[0027] The present invention provides an innovative genetic engineering-based method for improving the performance of filamentous fungal mycelial materials. By genetically modifying filamentous fungi, the present invention precisely regulates their chitin biosynthesis pathway, resulting in engineered bacteria with cell walls high in chitin. This strain, when used to produce mycelial materials, eliminates the need for exogenous chitin precursors to produce high-strength mycelial materials, overcoming the drawbacks of traditional induction methods and making it suitable for industrial production of mycelial materials. BRIEF DESCRIPTION OF THE DRAWINGS

[0028] Figure 1 This is the plasmid map of 1303-GLP-chitin in Example 1 of the present invention;

[0029] Figure 2 The plates were obtained 20 days after the transformation of the 1303-GLP-chitin plasmid in Example 1 of the present invention;

[0030] Figure 3 The standard curve of D-glucosamine content and chitin quality in Example 1 of the present invention;

[0031] Figure 4 Schematic diagram of the tensile testing machine used in Example 1 of the present invention for testing the strength of mycelium fiber skin.

[0032] The engineered bacteria GL-chintin-19 was classified as Ganoderma lucidum ( Ganoderma lingzhi ), deposited at China General Microbiology Culture Collection Center (CGMCC), deposited at Institute of Microbiology, Chinese Academy of Sciences, No. 3, Yard 1, Beichen West Road, Chaoyang District, Beijing, deposit number: CGMCC No. 42012, deposit date: May 21, 2025. DETAILED DESCRIPTION

[0033] In order to make the purpose, technical solutions and advantages of the present invention more clear, the present invention is further described in detail below with reference to specific embodiments. It should be understood that the specific embodiments described herein are only used to explain the present invention and are not intended to limit the present invention. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without making creative work are within the scope of protection of the present invention.

[0034] Mycelium, the fundamental unit of fungal materials, and chitin, a major component of the fungal mycelial cell wall, combine to create unique physical and chemical properties. Currently, mycelial materials are often prepared by exogenously adding chitin synthesis precursors during culture to induce the mycelium to produce more chitin, thereby increasing the strength of the mycelial material. However, this method increases process complexity and may lead to an uneven internal structure of the mycelium, resulting in a decrease in the mechanical properties of the mycelial material.

[0035] The present invention provides an engineered bacterium with a high chitin content. This engineered bacterium is constructed from a filamentous fungus through genetic modification. The genetic modification includes overexpression of chitin synthase, glutamine-fructose-6-phosphate transaminase, and uridine diphosphate-N-acetylglucosamine pyrophosphorylase. Under standard fermentation conditions, this engineered bacterium can increase chitin content in its cell walls without the addition of exogenous inducers such as chitin synthesis precursors, thereby improving the mechanical strength of the mycelial material obtained through fermentation.

[0036] An embodiment of the present invention further provides a method for improving the strength of a mycelium material, wherein the method comprises increasing the chitin, β-glucan, and a protein capable of binding to cell wall components in the mycelium cell wall.

[0037] The embodiment of the present invention also provides a Ganoderma lucidum engineered bacterium with high chitin content and a construction method thereof.

[0038] The embodiment of the present invention also provides an engineered bacterium GL-chintin-19 with a high chitin content.

[0039] The embodiments of the present invention also provide the use of the above-mentioned engineered bacteria, the engineered Ganoderma lucidum bacteria, and the engineered bacteria GL-chintin-19 in the production of mycelium materials.

[0040] The embodiment of the present invention also provides a method for producing mycelium fiber skin.

[0041] The solutions of the present invention are described below through specific embodiments.

[0042] The Ganoderma lucidum G10 used in the following examples is a commercial strain purchased from Wuhan Zhou Yulin Edible Fungi Research Institute.

[0043] The culture medium components used in the following examples are:

[0044] PDB medium (potato dextrose liquid medium): purchased from Tianjin Guangfu Technology Development Co., Ltd.;

[0045] CYM medium: maltose 10 g / L, glucose 20 g / L, tryptone 2 g / L, magnesium sulfate heptahydrate 0.5 g / L, potassium dihydrogen phosphate 4.6 g / L, yeast extract 2 g / L, low melting point agarose 15 g / L, vitamin B1 0.125 mg / L, mannitol 109.56 g / L;

[0046] Solid-state fermentation medium: sawdust 77.84% wt, bran 20% wt, sucrose 1% wt, gypsum 1% wt, potassium dihydrogen phosphate 0.1% wt, magnesium sulfate 0.05% wt, VB 0.01% wt.

[0047] Unless otherwise specified, the experimental methods in the following examples are conventional methods in the art and were performed according to the techniques or conditions described in the literature in the art or according to the product instructions. The materials and reagents used in the following examples were obtained from commercial sources unless otherwise specified.

[0048] Example 1

[0049] This embodiment provides a method for constructing an engineered Ganoderma lucidum strain with high chitin content.

[0050] 1. Plasmid construction

[0051] 1.1 PCR amplification

[0052] The endogenous CHS-5, GFAT, and UAP1 gene fragments of Ganoderma lucidum G10 were amplified by PCR using primers CHS-F / R (shown in SEQ ID NOs. 5 and 6), GFAT-F / R (shown in SEQ ID NOs. 7 and 8), and UAP-F / R (shown in SEQ ID NOs. 9 and 10) in Table 1. Overlap PCR was used to link these three gene fragments to form a CHS-linker-GFAT-linker-UAP gene fragment. The plasmid backbone 1303-GLP-iEGFP (purchased from Miaoling Biotechnology, formerly known as pCAMBIA1303-GpdA-3×FLAG-EGFP-TrpC-Hygro) was amplified by PCR using primers 1303-GLP-F / R (shown in SEQ ID NOs. 11 and 12).

[0053] Table 1 Primers and their sequences required for this experiment

[0054]

[0055] The reaction system and amplification conditions of the above PCR amplification are shown in Tables 2 and 3.

[0056] Table 2 PCR amplification reaction system

[0057]

[0058] Table 3 PCR amplification reaction conditions

[0059]

[0060] 1.2 Homologous recombination of multiple DNA fragments

[0061] The amplified product of the plasmid backbone 1303-GLP-iEGFP was ligated to the CHS-linker-GFAT-linker-UAP gene fragment using the ClonExpress Ultra One Step Cloning Kit (Nanjing Novozymes). The specific steps are as follows:

[0062] (1) Prepare the multi-fragment DNA homologous recombination reaction system as shown in Table 4 in a microcentrifuge tube with a total volume of 10 μL.

[0063] Table 4 Multi-fragment DNA homologous recombination reaction system

[0064]

[0065] (2) Mix thoroughly by shaking the bottom of the centrifuge tube.

[0066] (3) Incubate at 50°C for 15 min and immediately cool in crushed ice.

[0067] The gene expression pattern of recombinant plasmid 1303-GLP-chitin is as follows Figure 1 shown.

[0068] 2. Transformation and screening

[0069] 2.1 Protoplast preparation

[0070] Prepare a 2% (w / v) lytic enzyme solution and sterilize by filtration through a 0.22 μm microporous filter. Inoculate Ganoderma lucidum G10 in PDB medium and incubate at 25°C for 7 days. Mycelia are then picked and washed three times with 0.6 M mannitol (osmotic pressure stabilizer) and centrifuged at 6000 rpm for 10 minutes. The supernatant is discarded. Add 2% (w / v) lytic enzyme solution to 0.2 g of wet mycelia per 1 mL of enzyme solution. Vortex thoroughly to break up the mycelia. Incubate in a 31°C waterbath for 3.5 hours. Gently vortex every 0.5 hours during the enzymatic digestion. After the digestion is complete, remove mycelial fragments through a 0.45 μm microporous filter. Centrifuge the filtrate at 4000 rpm for 10 minutes, and discard the supernatant to obtain the protoplast pellet. Redissolve the protoplast pellet in 0.6 M mannitol and centrifuge at 4000 rpm for 10 minutes. Repeat three times to completely remove the enzyme solution. The protoplasts were added to an appropriate amount of 0.6 M mannitol solution to prepare a protoplast suspension.

[0071] The above experimental procedure was repeated three times, and the number of protoplasts in each ml of suspension was calculated using a hemocytometer.

[0072] 2.2 Protoplast transformation

[0073] The 1303-GLP-chitin recombinant plasmid was combined with the protoplasts of the mononuclear sessile Ganoderma lucidum strain (10 7 PEG-mediated transformation:

[0074] The protoplast suspension was centrifuged at 1258×g, 4°C for 5 min, the supernatant was discarded, and STC buffer (sorbitol 218.6 g / L, 0.01M Tris-HCl, CaCl2 11.1 g / L) was added and gently mixed to 100 μL to obtain the STC suspension of protoplasts; 30 μg of 1303-GLP-chitin recombinant plasmid, 100 μg of heparin sodium, and 0.25 mM spermidine were mixed with water to 100 μL, added to the STC suspension of protoplasts, mixed quickly, and placed on ice for 10 min. Add 200 μL of PTC (75% PEG4000, 1 mM Tris-HCl (pH=7.5), 2.5 mM CaCl2), gently rotate the centrifuge tube to mix, place on ice for 10 min, add 200 μL of PTC, gently rotate the centrifuge tube to mix, place on ice for 10 min, add 800 μL of PTC, flick to mix, and incubate at 30°C for 30 min. Add 1 mL of STC buffer to dilute the transformation system, and finally add it to 20 mL of CYM medium (containing a final mass concentration of 4 mg / L of carboxin), mix well, and pour it into the plate. After the plate solidifies, place it in a 26°C constant temperature incubator and culture for 10-15 days. Visible colonies will grow. Culture until the 20th day ( Figure 2 Resistant colonies were transferred to fresh CYM medium (containing carboxin at a final concentration of 4 mg / L) for secondary screening and five consecutive subcultures to test their stability. A total of 35 stable mutants were obtained. Nine of these strains were randomly selected and the relative expression of the CHS gene was determined by qPCR. As shown in Table 5, due to the random integration of exogenous DNA by PEG transformation, different strains of Ganoderma lucidum expressed different levels of the CHS-5 gene.

[0075] Table 5 Relative expression of CHS-5 gene in resistant colonies

[0076]

[0077] From the positive transformants, G10-chitin-15 and G10-chitin-19 strains with relatively high expression of the CHS-5 gene were selected for subsequent experiments.

[0078] 3. Detection of chitin content in mutant strains

[0079] The monokaryotic strains G10, G10-chitin-15, and G10-chitin-19, with varying relative expression levels, were cultured in CYM liquid shake flasks. Mycelia were harvested after 6 days of culture, cleaned, and freeze-dried under vacuum. Different masses of chitin powder were weighed to create a standard curve. Chitin content was determined by weighing 0.01 g of freeze-dried mycelial powder from G10, G10-chitin-15, and G10-chitin-19:

[0080] 400 μL of 4 M HCl was added to 0.01 g of ground mycelium and heated at 96°C for 16 h. 400 μL of autoclaved water was then added to cool the reaction. 9 mg of ground activated carbon was added to each sample and vortexed three times within 1 hour. The samples were then centrifuged, and 300 μL of the supernatant was diluted 10-fold with 2.63 mL of autoclaved distilled water and 70 μL of 8 M aqueous NaOH to neutralize the samples. The relative percentage of chitin after extraction was determined using a D-glucosamine assay kit (Megazyme, Ireland) according to the manufacturer's protocol.

[0081] Obtain a standard curve of D-glucosamine content and chitin quality (such as Figure 3 The chitin contents of G10, G10-chitin-15, and G10-chitin-19 were calculated and shown in Table 6. This indicates that the chitin contents of the mutants obtained using the gene editing method of the present invention were increased by 110% and 548%, respectively, compared to the starting strains. Chitin content in these strains increased with increasing integrated copy number and relative expression level.

[0082] Table 6 Chitin content determination of mutant strains

[0083]

[0084] The G10-chitin-19 strain was deposited on May 21, 2025, at the General Microbiology Center of the China Culture Collection Administration, No. 3, Yard 1, Beichen West Road, Chaoyang District, Beijing, with the deposit number CGMCC No. 42012 and named GL-chintin-19.

[0085] Example 2

[0086] This example provides the use of the monokaryotic strain G10 and G10-chitin-15 and G10-chitin-19 constructed in Example 1 in the production of mycelial materials.

[0087] Liquid inoculum of monokaryotic strains G10, G10-chitin-15, and G10-chitin-19 (5 g of mycelia each) (cultured in shake flasks under the same conditions and filtered to obtain mycelia) was solid-state fermented in a solid-state fermentation medium to obtain mycelial fiber skins. The specific fermentation method is as follows:

[0088] 1. Add water to the solid-state fermentation medium to adjust the moisture content to 65% wt as the fermentation base;

[0089] 2. Dispense the above base materials into 2400ml culture boxes according to a 50% filling factor and cover the box lid;

[0090] 3. Stack the culture boxes evenly in the autoclave and sterilize at 121°C for 90 minutes.

[0091] 4. After sterilization, cool the culture box to room temperature in a clean room;

[0092] 5. In a clean bench, open the lid of the box and evenly spray 70 mL of liquid inoculum (containing 5 g of mycelium) on the surface of the base material and close the lid;

[0093] 6. Place the culture room in a constant temperature room at 28-30°C and RH 60%-70% for 15 days;

[0094] 7. After the cultivation cycle is over, open the lid of the box and separate and remove the fungus skin above the substrate.

[0095] Cut the mycelium sheet of 2.5 cm × 10 cm in size and fix it on the tensile testing machine to test the strength of the mycelium sheet (such as Figure 4The results are shown in Table 7. Compared with the starting strain G10, the strength of the mycelial materials obtained by G10-chitin-15 and G10-chitin-19 using the same method increased by 78% and 149% respectively.

[0096] Table 7 Mutant mycelium fiber skin strength test

[0097]

[0098] These results indicate that G10-chitin-15 and G10-chitin-19 can produce high-strength mycelial materials under conventional fermentation conditions without exogenous addition of chitin precursors, and have the potential for industrial application in producing high-strength mycelial materials.

[0099] The above description is only a preferred embodiment of the present invention and is not intended to limit the present invention. Any modifications, equivalent substitutions or improvements made within the spirit and principles of the present invention should be included in the scope of protection of the present invention.

Claims

1. An engineered bacterium, characterized in that: The engineered bacteria is Ganoderma lucidum ( Ganoderma lingzhi ) is a starting strain, which is constructed by genetic modification; the genetic modification includes: overexpressing chitin synthase, glutamine-fructose-6-phosphate transaminase, and uridine diphosphate-N-acetylglucosamine pyrophosphorylase; the chitin synthase is chitin synthase CHS-5, the amino acid sequence of which is shown in GenBank: AZQ26802.1; the uridine diphosphate-N-acetylglucosamine pyrophosphorylase is uridine diphosphate-N-acetylglucosamine pyrophosphorylase UAP1, the amino acid sequence of which is shown in SEQ ID NO.2; and the glutamine-fructose-6-phosphate transaminase is GFAT, the amino acid sequence of which is shown in SEQ ID NO.

1.

2. The engineered bacteria according to claim 1, characterized in that The method for overexpressing the chitin synthase, glutamine-fructose-6-phosphate transaminase and uridine diphosphate-N-acetylglucosamine pyrophosphorylase is to increase the copy number of the coding gene of each enzyme.

3. The engineered bacteria according to claim 1, characterized in that The nucleotide sequence of the gene encoding the chitin synthase CHS-5 is shown in GenBank: MH463055.1, the nucleotide sequence of the gene encoding the glutamine-fructose-6-phosphate transaminase GFAT is shown in SEQ ID NO.3, and the nucleotide sequence of the gene encoding the uridine diphosphate-N-acetylglucosamine pyrophosphorylase UAP1 is shown in SEQ ID NO.

4.

4. An engineered strain GL-chintin-19, characterized in that: Its classification name is Ganoderma lucidum ( Ganoderma lingzhi ), deposited on May 21, 2025, at the General Microbiology Center of China Culture Collection Administration, address: No. 3, Yard 1, Beichen West Road, Chaoyang District, Beijing, with the deposit number CGMCC No. 42012.

5. Use of the engineered bacteria according to any one of claims 1 to 3 or the engineered bacteria GL-chintin-19 according to claim 4 in the production of mycelium materials.

6. A method for producing mycelium fiber skin, characterized in that: The mycelium fiber skin is obtained by solid-state fermentation using the engineered bacteria according to any one of claims 1 to 3 or the engineered bacteria GL-chintin-19 according to claim 4.

7. The method according to claim 6, characterized in that The mycelium fiber skin is obtained by solid-state fermentation of the engineered bacteria GL-chintin-19 according to claim 6.

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