Bidirectional promoter derived from thermophilic fungus destructor thermophilus and application thereof

Abstract

The invention relates to the technical field of agricultural biology, in particular to a bidirectional promoter derived from thermophilic fungus destructor thermophilus and application of the bidirectional promoter. The bidirectional promoter derived from thermophilic fungus destructor thermophilus has unique bidirectional transcriptional activity characteristics, and can simultaneously and efficiently start transcriptional expression of two independent genes in opposite directions in the same regulatory region. A more favorable promoter element is provided for metabolic engineering modification of industrial filamentous fungus myceliophthora thermophila, the construction process of a multi-gene co-expression system is remarkably simplified, and synergistic expression of two functional genes can be realized through a single promoter element; the method has wide application space and market prospect in production of enzyme preparations, biological medicine products and the like.

Description

Technical Field

[0001] The present invention relates to the field of agricultural biotechnology, and in particular to a bidirectional promoter derived from the thermophilic fungus Myceliophthora thermophila and applications thereof. Background Art

[0002] Myceliophthora thermophila is a high-temperature filamentous fungal chassis strain with the ability to efficiently degrade biomass and secrete proteins. No viruses, mycoplasmas, or reported fungal toxins have been detected. Compared with traditional industrial strains Trichoderma reesei and Penicillium decumbentum, the cellulase produced by Myceliophthora thermophila has significantly higher specific activity and thermal stability. Its enzyme preparations perform particularly well in high-temperature industrial environments and have important application value in biofuels, textiles, papermaking and other fields. At the same time, the glycosylation modification system of this strain can produce human-like glycan structures. This feature gives it a unique advantage in the production of recombinant protein drugs and vaccines. Therefore, the transformation of Myceliophthora thermophila through genetic engineering technology and its development as a multifunctional filamentous fungal chassis cell is of great significance for the production of enzyme preparations and biopharmaceutical products.

[0003] Currently, research on the gene expression regulation system of Myceliophthora thermophila is still at a relatively early stage. The number of functional promoter elements identified in this strain is relatively limited, with only two widely used for gene expression regulation: the transcription elongation factor EF-1 promoter and the pyruvate decarboxylase PDC promoter. This scarcity of promoter resources has become a key bottleneck in complex metabolic engineering projects requiring multi-gene co-expression, severely limiting the potential of Myceliophthora thermophila for applications in synthetic biology and metabolic engineering. Therefore, developing novel multifunctional promoters (such as bidirectional promoters and conditionally inducible promoters) through genome mining, rational design, or directed evolution, and establishing a comprehensive toolbox for gene expression regulation, will be a key research direction to overcome the bottleneck of metabolic engineering in Myceliophthora thermophila and hold important theoretical and practical significance for its industrial application.

[0004] Bidirectional promoters, as specialized cis-acting elements, are located within the shared DNA sequence region between two oppositely aligned genes in the genome. Their structure enables RNA polymerase to initiate transcription simultaneously from opposite directions, effectively coordinating the co-expression regulation of two independent genes. Existing studies have shown that, although bidirectional promoters from Pichia pastoris have been successfully identified, these promoters exhibit atypical promoter characteristics: they lack the TATA box core domain, a universally conserved structure in eukaryotic promoters; sequence homology varies widely between different bidirectional promoters; and their functional activity exhibits species-specific characteristics. Consequently, traditional bioinformatics prediction methods based on sequence conservation analysis (such as motif searching and promoter prediction) have limitations, making it difficult to effectively identify functional bidirectional promoter sequences in Myceliophthora thermophila using conventional computational biology methods. Summary of the Invention

[0005] The purpose of the present invention is to provide a bidirectional promoter derived from the thermophilic fungus Myceliophthora thermophila.

[0006] Another object of the present invention is to provide the application of the bidirectional promoter derived from the thermophilic fungus Myceliophthora thermophila.

[0007] The present invention provides bidirectional promoters BIP1, BIP2, BIP3, BIP4, BIP5 and BIP6 derived from the thermophilic fungus Myceliophthora thermophila. The nucleotide sequences of the bidirectional promoters are shown in SEQ ID NO: 1, SEQ ID NO: 2, SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 5 and SEQ ID NO: 6.

[0008] SEQ ID NO: 1: TGCGACGGTGCTGAGGTTTGTCGAAGAAGTGAGACTCAAATCGGCTCCTGGCGGTGTTGTCCTCGTCGCTGGCTGGGGTTGTCGCTGAGGGAGCAGAATCGCAAACTCGCCGAAATATTCTTTTTGCCGATGTGAATTCGCCTGCAGAGTGGCTGAGTTCTCATTGGCTGAGCTTTGGGTGTGCCAGAG GTTCTCCGAAACCCTAACGGCTCCACTCTTTTCCGACCAACAACTCGGCCGGTTGCATGACGAATCTTTGGATTGGTAATCGCGAGGCGAATTTGATTTGGTCAAAACCGCAGAGGCTCCCAACCATCCCCTCTCCCCAAGACCAAGTCGATACCCGTTGTCGTGACATCGCAAAAACACCGTCACA.

[0009] SEQ ID NO: 2: TGTGAGAGATATCGGTGTCGAGTCAAATGGGCGATGGGCGACGATTCCTTTGACAGCGACGGGGTCTTCGGGTCGTTGGAAGGGAATAGCCTCTCCTGCCGGCACAACGTCCTAACAAAATGTGAGAGGTGTGGATGCGCAAAATGCCAGCCAGGGCGGAAAACCCTACAGCCGAAAGTGGCCCGTGCAGCACAATCTTGGTCCAGCCATGATGGTCCCGCAGTTCCGAAAGCGGTTAGCTTGGGGTTTGGCCTTTCCGAACTTGCGCACAACCGAAGGCTCGTCCATTTCAAAATCTTGAGCAGCAGCAAGTCAAGTTGACGACCGACCAACGAATTTACTAATCCACCAAACAGCCAAAGCGCTCAAG。

[0010] SEQ ID NO:3: CTTGTCGTTGAGGCTTTCAGAAGTTGGTCGACCGAGCTGAAATTCGCAAAGTTGCAACGGGGCCCTGAAGGTTCGAGGAAAATTGAGGCACGAGGGGGATTTGTGGTGGTCCGTGCGTATCGCTCCCGCGCGTGCCCTAACGATAGCCTGCTGGTCCAAGCACCTGCACCCTGCTTAGTCACCCAGCCCGAAATCCCGTCTTGGATTTGCTTCCACACAAAAATACCCCAATTTTTTCGCAGCCCGCAGCAACTAACGACTTCGACACATCGATCCACGCCGACACACCCAGTAAACAGGCAAG。

[0011] SEQ ID NO:4: TTTGGCGGTTGTTGGCCGATCGGGGCGAGATTCTCAATTCGTAAAGAGGAAGAGAGGTATCTACCAGCATGCCGAAGCGTGTGACGTTAGCCTTCCGAGGCTCGATGAGCGTGGAGAGTGCCCTCGTCGACAAGTCGAGAGGGTTCTGCCTGCTCAGTTGCCGACGAGGAGAAGCGTATCACCGGAGGGCGTCGGCCACTGCCTCAGCCATCCCGTGATTGCTCTCCCCGGCCTTCGCTGAAGGCATTTGACTGTTGACAAGTCATTTTTCAGAAAACCACATACCTCAGGTTTGCCTGCGTTGAGAAGCCGTGATGGTTGATCAGGTCCTGGATCGAGGAGGTTTGGTTGCGACCAGAAAATTCAGCAGGTGTTGAGCGAGCTGTGGTGGCCCGTGCCCCGGAAACTTGCCCTAAACGCTTGGGGTCCCAGATTTGTGCGCTTGCTCAAAGCAGACTAGCTCGGTGTGGAATCCTTTCGACCACACAAAATCCGCGAATTGAGGTCCTGGGTCGGGTCCGTGGGGCCCGACAGCCCGCACACTTTAAACTTGACTTTTGTGGCCCAGCCGAGATCCCTCCATCCAACACACGAAAAACGACTACACTATCACCTACTCTTCGAGGATCGTAGCCGACACTTACCAGCAAACAACAACGTCAAG。

[0012] SEQ ID NO:5: TTTGGCGGTGTCTCGAGTATGCTGGTTCTCAAGAGGTTGTGTGGGAGAAGGGCGAGCAGGTCAGAAATTCGTCTGCTGCTGGAAAACTGCCGTGTTGGACCCACCTGAGAAATTCCCCACCCTCAATCAGCTCTTGGGGTGCGACTGAGGGCAAGTTCTCGAGCAAACCCTAAACCGCAGGCACAGAGTCAGCTGACCCACATCACCCAGAGTGGGCCCCGCCCAAAAGTTCAACTCCGCCGCCGACGCACGTCAATTTCCTGCCATTTTCTTCCCAAGCGCACCTTCACAACATCTACAGATTGCTGAGGAAGACATTGTGTCTAGGTACGTCTGCACGATGTTGCGAGCCGGCGCTTCGTCCCTTTTGCCTTTTGAGGCACCATGATACCCAATCTTGTCATCTCTTCTGCTGCGGGCTGCTATGGAATCTTCGCTCGCGGCTCGGACACTTGCCCGAGCGCGGTTTCGAGTCGGCTTCCCCCCTCCCCTCCCCATGATCCAAGATGGCGCCGGCTTAGCTCTTCACGAAGCCTTCTCTCCCCTGCCTCTAACTCATCCTCACCACTTCTCAGTAGGCAATC。

[0013] SEQ ID NO:6: TTTGATGGATGTTCTGGTGGTTGATGAAGGTTGGAGTCGGGGTCGAGGCGCAAAAGGCGGGGAGTTTTTTCTGAGAGGCCGCGCTGCCAGATGCCAAAGCTTAGGTTAACCGAGCCCTAATAAAATGGGGCGCTCTGAGTGGCTGCGCTCCCGCCCAGACCACTCCCAGCGGGGCCACCAAAAAATTCACGACCACACGCTCAAGCCTTCTGTGTCGTCGTCCTGCACGCCAAACTTCCAATCCATCACCACCTTCAGCTTTTCAGCCAGCCGACCACCGCAAAC。

[0014] The present invention also relates to a recombinant expression vector comprising the above-mentioned bidirectional promoter derived from the thermophilic fungus Myceliophthora thermophila.

[0015] The beneficial effects of the present invention are that the bidirectional promoters BIP1, BIP2, BIP3, BIP4, BIP5, and BIP6, derived from the thermophilic fungus Myceliophthora thermophila, possess unique bidirectional transcriptional activity characteristics, capable of efficiently and simultaneously initiating the transcriptional expression of two independent genes in opposite directions within the same regulatory region. This invention provides more favorable promoter elements for metabolic engineering of the industrial filamentous fungus Myceliophthora thermophila, significantly simplifying the construction process of a multigene co-expression system. A single promoter element can achieve the coordinated expression of two functional genes, demonstrating broad application and market prospects in the production of enzyme preparations, biopharmaceuticals, and the like. BRIEF DESCRIPTION OF THE DRAWINGS

[0016] Figure 1 A schematic diagram of a recombinant expression vector containing a bidirectional promoter derived from the thermophilic fungus Myceliophthora thermophila is shown; Figure 2 It shows the transcription of the green fluorescent gene from the bidirectional promoter of the thermophilic fungus Myceliophthora thermophila; Figure 3 It shows the transcription of the red fluorescent gene from the bidirectional promoter of the thermophilic fungus Myceliophthora thermophila; Figure 4 It shows the expression of green fluorescent protein from the bidirectional promoter of the thermophilic fungus Myceliophthora thermophila; Figure 5 Shows the expression of red fluorescent protein from the bidirectional promoter of the thermophilic fungus Myceliophthora thermophila. DETAILED DESCRIPTION

[0017] The present invention is further illustrated by way of examples below, but the present invention is not limited to the scope of the examples. Experimental methods in the following examples where specific conditions are not specified were performed according to conventional methods and conditions, or selected according to the product specifications.

[0018] 1. Strains Myceliophthora thermophila ATCC42464 is a thermophilic filamentous fungus currently used for the production of industrial enzyme preparations and is stored in the American Type Culture Collection. ku 80 was used as the starting strain. Compared with Myceliophthora thermophila ATCC42464, strain Δ ku 80 has lost its nonhomologous DNA repair function, a property that makes it suitable for site-specific gene integration and maintenance of equal gene copy number.

[0019] 2. Culture medium and other reagents The culture medium used for culturing Myceliophthora thermophila was potato dextrose agar medium; Protoplast regeneration medium (100 mL): 2 mL 50 × Vogel's solution, 2 g sucrose, 18.2 g sorbitol, 0.75 g agarose, dilute to 100 mL; 50 × Vogel's solution (1 L): 130 g trisodium citrate dihydrate, 126 g potassium nitrate, 144 g ammonium dihydrogen phosphate, 80 g potassium dihydrogen phosphate, 10 g magnesium sulfate heptahydrate, 5 g calcium chloride dihydrate, 5 mL trace element solution, 2.5 mL 0.1 mg / mL biotin solution, 2 mL chloroform, dilute to 1000 mL; Trace element solution (100 mL): 5 g citric acid monohydrate, 5 g zinc sulfate heptahydrate, 1 g ferrous ammonium sulfate hexahydrate, 0.25 g copper sulfate pentahydrate, 0.05 g manganese sulfate monohydrate, 0.05 g boric acid, 0.05 g sodium molybdate dihydrate, dilute to 100 mL; Functional evaluation medium (100 mL): 7.5 g glucose, 1 g yeast extract, 0.015 g potassium dihydrogen phosphate, 0.015 g potassium hydrogen phosphate, 0.01 g magnesium sulfate heptahydrate, 0.01 g calcium chloride dihydrate, 0.1 mL 0.1 mg / mL biotin solution, 0.1 mL trace element solution.

[0020] This study used whole-genome sequencing to locate and analyze the coding genes of Myceliophthora thermophila. By systematically identifying intergenic regions, candidate non-coding regulatory regions were identified. Furthermore, combined with RNA-seq transcriptome data from cultures grown under different carbon sources (glucose / cellulose), and based on colocalization analysis of differentially expressed genes, short promoter sequences with potential bidirectional transcriptional regulation were identified.

[0021] The present invention provides bidirectional promoters BIP1, BIP2, BIP3, BIP4, BIP5 and BIP6 derived from the thermophilic fungus Myceliophthora thermophila. The nucleotide sequences of the bidirectional promoters are shown in SEQ ID NO: 1, SEQ ID NO: 2, SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 5 and SEQ ID NO: 6. Example 1 Preparation of a recombinant expression vector containing a bidirectional promoter derived from the thermophilic fungus Myceliophthora thermophila

[0022] PCR amplification was performed using the genomic DNA of Myceliophthora thermophila as a template, and electrophoresis was performed on a 1% agarose gel. The target fragment, i.e., the endogenous bidirectional promoter of Myceliophthora thermophila, was recovered from the gel. Subsequently, the fragment was recovered and connected to the HygR-Vector-EGFP-BIP-DsRed vector backbone by homologous recombination to obtain the recombinant expression vectors HygR-Vector-EGFP-BIP1-DsRed, HygR-Vector-EGFP-BIP2-DsRed, HygR-Vector-EGFP-BIP3-DsRed, HygR-Vector-EGFP-BIP4-DsRed, HygR-Vector-EGFP-BIP5-DsRed and HygR-Vector-EGFP-BIP6-DsRed. The primers and sequences used for PCR amplification of the bidirectional promoter are shown in Table 1 (the italicized part is the homology arm). The recombinant expression vector containing the bidirectional promoter from the thermophilic fungus Myceliophthora thermophila is shown in Table 1. Figure 1 shown.

[0023] Table 1 Primer information for constructing bidirectional promoter recombinant expression vector Primer Name Sequence Information (5'-3') Vector-EGFP-BIP-DsRed-F (SEQ ID NO:7) ATGGACAACACCGAGGACGTCATC Vector-EGFP-BIP-DsRed-R (SEQ ID NO:8) ATGGTGAGCAAGGGCGAGGAGCTG BIP1-F (SEQ ID NO:9) TTGCGACGGTGCTGAGGTTTG BIP1-R (SEQ ID NO:10) TGTGACGGTGTTTTTGCGAT BIP2-F (SEQ ID NO:11) TTGTGAGAGATATCGGTGTCG BIP2-R (SEQ ID NO:12) CTTGAGCGCTTTGGCTGTTTG BIP3-F (SEQ ID NO:13) TCTTGTCGTTGAGGCTTTCAGAAG BIP3-R (SEQ ID NO:14) CTTGCCTGTTTACTGGGTGTG BIP4-F (SEQ ID NO:15) TTTTGGCGGTTGTTGGCCGATCG BIP4-R (SEQ ID NO:16) CTTGACGTTGTTGTTTGCTGG BIP5-F (SEQ ID NO:17) TTTTGGCGGTGTCTCGAGTATGC BIP5-R (SEQ ID NO:18) GATTGCCTACTGAGAAGTGGTG BIP6-F (SEQ ID NO:19) TTTTGATGGATGTTCTGGTGGTTG BIP6-R (SEQ ID NO:20) GTTTGCGGTGGTCGGCTGGCTG . Example 2 Construction of an engineered strain of Myceliophthora thermophila overexpressing a bidirectional promoter recombinant expression vector

[0024] Protoplast preparation: Thermomycete ku 80 spore suspensions were spread on potato dextrose agar plates and cultured at 37 °C for 16 h. Germinated mycelia were collected and placed in 20 mL of 10 mg / mL lytic enzyme solution and digested at 75 rpm at 28 °C for 2 h. The filtrate was collected by filtration into a 50 mL centrifuge tube and centrifuged at 3000 rpm at 4°C for 10 minutes. Then, 10 mL of STC solution (1.2 M sorbitol, 50 mM calcium chloride, 35 mM sodium chloride, pH 7.0) was added to suspend the protoplasts and centrifuged at 3000 rpm at 4°C for 10 minutes. The supernatant was discarded and 200 μL of STC solution was added to resuspend the protoplasts. 50 μL of PEG solution and the gene fragment were gently shaken and incubated on ice for 20 minutes. Then, 2 mL of PEG solution was added and gently shaken to mix. The tube was incubated at room temperature for 5 minutes. 4 mL of STC solution was added and then mixed with protoplast regeneration medium supplemented with 100 μg / mL hygromycin. The tube was cultured at 45°C for 3 days until transformants grew.

[0025] The transformants were picked and cultured on potato dextrose agar plates for 3 days, and then the genomic DNA of the transformants was extracted and used as a template for PCR verification. ku80 was used as the host, and the gene fragments of the recombinant vectors HygR-Vector-EGFP-BIP1-DsRed, HygR-Vector-EGFP-BIP2-DsRed, HygR-Vector-EGFP-BIP3-DsRed, HygR-Vector-EGFP-BIP4-DsRed, HygR-Vector-EGFP-BIP5-DsRed, and HygR-Vector-EGFP-BIP6-DsRed were transformed to obtain the thermophilic myceliophthora engineering strain Δ containing the bidirectional promoter recombinant expression vector ku 80-EGFP-BIP1-DsRed, Δ ku 80-EGFP-BIP2-DsRed, Δ ku 80-EGFP-BIP3-DsRed, Δ ku 80-EGFP-BIP4-DsRed, Δ ku 80-EGFP-BIP5-DsRed and Δ ku 80-EGFP-BIP6-DsRed. Example 3 Quantitative evaluation of transcription efficiency of bidirectional promoters

[0026] Thermophilic Myceliophthora starting strain Δ ku 80. The thermophilic Myceliophthora engineering strain overexpressing the bidirectional promoter recombinant expression vector was inoculated on a potato dextrose agar plate and cultured at 45°C for 7 days until spores were produced. The spore suspension was washed and then cultured according to 10 6 The spores were inoculated into functional evaluation medium at a concentration of 10 spores / mL. After 3 days of fermentation at 45°C, the cells were harvested and RNA was extracted using the Trizol method. The expression levels of EGFP and DsRed genes were determined by quantitative PCR. An engineered strain of Myceliophthora thermophila overexpressing a recombinant expression vector containing a fusion bidirectional promoter (the fusion promoter was constructed by reverse-linking the two constitutively high-expressing promoters of Myceliophthora thermophila, P87219 and P2074790) was used as a positive control to compare transcriptional differences between strains. The primers and sequences used for quantitative PCR are shown in Table 2.

[0027] Table 2 Fluorescence quantitative PCR primer information Primer Name Sequence Information (5'-3') Actin-F (SEQ ID NO:21) TGCCCATCTACGAGGGTTTC Actin-R (SEQ ID NO:22) TTGATGTCACGGACAATTTCAC EGFP-F (SEQ ID NO:23) GACTGGGTGCTCAGGTAGTG EGFP-R (SEQ ID NO:24) CAAGATCCGCCACAACATCG DsRed-F (SEQ ID NO:25) GACCGTGTACAAGGCCAAGA DsRed-R (SEQ ID NO:26) AGTCCTCGTTGTGGTTGGTG .

[0028] The results are as follows Figure 2 and Figure 3 As shown, compared with the thermophilic myceliophthora engineering strain overexpressing the recombinant expression vector fused with a bidirectional promoter, the thermophilic myceliophthora engineering strain overexpressing the bidirectional promoter recombinant expression vector Δ ku 80-EGFP-BIP1-DsRed, Δ ku80-EGFP-BIP2-DsRed, Δ ku 80-EGFP-BIP3-DsRed, Δ ku 80-EGFP-BIP4-DsRed, Δ ku 80-EGFP-BIP5-DsRed and Δ ku The expression levels of green fluorescence and red fluorescence genes of 80-EGFP-BIP6-DsRed were significantly higher than those of the positive strains, and the thermophilic myceliophthora starting strain Δ ku No green fluorescent and red fluorescent gene expression was detected in 80, indicating that the bidirectional promoters BIP1, BIP2, BIP3, BIP4, BIP5 and BIP6 derived from the thermophilic fungus Myceliophthora thermophila have efficient bidirectional transcriptional activity characteristics and can efficiently initiate the transcriptional expression of two independent genes in opposite directions in the same regulatory region. Example 4 Qualitative evaluation of bidirectional promoter-driven protein expression

[0029] Thermophilic Myceliophthora starting strain Δ ku 80. The engineered strain of Myceliophthora thermophila overexpressing the recombinant expression vector of the bidirectional promoter was inoculated on a potato dextrose agar plate and cultured at 45°C for 7 days until spores were produced. The spore suspension was washed and quantified to 10 8 spores / mL, and after thorough mixing, 200 μL of the spore suspension was transferred to a Corning® 96-well black polyester microplate. Spore fluorescence intensity was measured using a BioTek SYNERGY H1 multi-function microplate reader. Parameters were: gain 100; detection height 7 mm; excitation and emission wavelengths of 488 nm and 520 nm for green fluorescent protein (EGFP); and 558 nm and 585 nm for red fluorescent protein (DsRed).

[0030] The results are as follows Figure 4 and Figure 5 As shown, the starting strain of Myceliophthora thermophila Δ ku Compared with 80, the thermophilic myceliophthora engineering strain overexpressing the bidirectional promoter recombinant expression vector Δ ku 80-EGFP-BIP1-DsRed, Δ ku 80-EGFP-BIP2-DsRed, Δ ku 80-EGFP-BIP3-DsRed, Δ ku 80-EGFP-BIP4-DsRed, Δ ku 80-EGFP-BIP5-DsRed and Δ kuThe green fluorescence value and red fluorescence value of 80-EGFP-BIP6-DsRed were significantly enhanced, indicating that the bidirectional promoters BIP1, BIP2, BIP3, BIP4, BIP5 and BIP6 derived from the thermophilic fungus Myceliophthora thermophila can efficiently drive the expression of proteins encoded by two independent genes in opposite directions simultaneously within the same regulatory region.

[0031] The above embodiments are only used to explain the technical solutions of the present application and do not limit the scope of protection of the present application.

Claims

1. A bidirectional promoter of the thermophilic fungus Myceliophthora thermophila, characterized in that The nucleotide sequence of the bidirectional promoter is shown in SEQ ID NO: 1, SEQ ID NO: 2, SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 5 or SEQ ID NO:

6.

2. A method for bidirectional gene expression in Myceliophthora thermophila, characterized in that: The method comprises the following steps: Constructing a recombinant expression vector comprising a bidirectional promoter and a target gene, wherein the nucleotide sequence of the bidirectional promoter is shown in SEQ ID NO: 1, SEQ ID NO: 2, SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 5 or SEQ ID NO: 6; The obtained recombinant expression vector was introduced into Myceliophthora thermophila cells.

3. Use of the bidirectional promoter of the thermophilic fungus Myceliophthora thermophila according to claim 1 for bidirectionally promoting gene expression in Myceliophthora thermophila. 4 . A recombinant expression vector comprising the bidirectional promoter of the thermophilic fungus Myceliophthora thermophila according to claim 1 .

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