Recombinant transdermal type VII collagen as well as preparation method and application thereof

Abstract

The invention belongs to the technical field of protein engineering and genetic engineering, and particularly relates to recombinant transdermal VII collagen white and a preparation method and application thereof. Specifically, the recombinant transdermal VII-type collagen provided by the invention is obtained by preferably selecting a sequence containing 54 amino acids, connecting 6 copies in series and introducing a cell-penetrating peptide to an N terminal, and tests prove that compared with commercially available recombinant VII-type collagen, the recombinant transdermal VII-type collagen has better transdermal performance and cell proliferation promoting performance, and can be used for preparing a recombinant VII-type collagen. Due to the good transdermal property, the hydrogel can enter the epidermis to play a role, and healing of epidermis wounds is promoted; meanwhile, the preparation method of the recombinant transdermal type VII collagen is simple and easy to implement, and is very suitable for industrial mass production, so that the recombinant transdermal type VII collagen has a good practical application value.

Description

Technical Field

[0001] This invention belongs to the fields of protein engineering and genetic engineering technology, specifically relating to a recombinant transdermal type VII collagen, its preparation method, and its application. Background Technology

[0002] The information disclosed in this background section is intended only to enhance understanding of the overall background of the invention and is not necessarily to be construed as an admission or in any way implying that such information constitutes prior art known to those skilled in the art.

[0003] Collagen, the most abundant structural protein in the human body, is widely distributed in tissues such as skin, bones, tendons, and cartilage, and is a core component for maintaining the integrity of tissue structure and physiological function. Among them, type VII collagen (COL7), as the main structural protein anchoring fibers, is located in the dermal-epidermal junction (DEJ) of the skin. Through interactions with molecules such as laminin and integrins, it constructs the mechanical connection between the epidermis and dermis, playing an irreplaceable role in maintaining the skin's barrier function, elasticity, and resistance to damage. Furthermore, the functional value of type VII collagen is particularly prominent in skin physiology and repair processes. Therefore, exogenous supplementation of type VII collagen has become an important way to improve skin condition and promote skin repair.

[0004] Based on the aforementioned application needs, the preparation and application of recombinant type VII collagen has become a research hotspot in the fields of bioengineering, daily chemical products, and biomedical materials. Currently, various expression systems for recombinant type VII collagen have been developed, including prokaryotic expression systems (such as *E. coli*) and eukaryotic expression systems (such as yeast and mammalian cells). However, the inventors have discovered that recombinant type VII collagen prepared using existing technologies still has significant technical defects: First, natural collagen has a large molecular weight and strong hydrophilicity, making it difficult to penetrate the stratum corneum barrier of the skin. When supplemented exogenously, it mostly remains on the skin surface and cannot penetrate into the deep epidermis or even the dermis to exert its effects, greatly limiting its application in skin repair, anti-aging, and other scenarios. Second, the function of type VII collagen depends on its specific triple helix structure and active site. Existing recombinant protein sequence designs often directly extract natural sequence fragments, leading to some recombinant proteins folding incorrectly and active sites being masked, thus affecting its biological functions such as promoting cell proliferation and mediating cell adhesion. In addition, some recombinant expression systems have problems such as low expression levels, difficulty in controlling product purity, and poor product stability during large-scale fermentation, resulting in high production costs and difficulty in meeting the needs of industrial applications. At the same time, if overly complex processes are used in the separation and purification process, the natural structure of collagen may be destroyed, further reducing its biological activity. Summary of the Invention

[0005] To address the shortcomings of existing technologies, the present invention aims to provide a recombinant transdermal type VII collagen, its preparation method, and its applications. The recombinant transdermal type VII collagen prepared by this invention exhibits excellent transdermal properties and cell proliferation-promoting properties. Furthermore, this invention provides a method for preparing recombinant collagen, which is highly suitable for large-scale industrial production. Based on the above research results, this invention has been completed.

[0006] To achieve the above-mentioned technical objectives, the present invention provides the following technical solution: A first aspect of the present invention provides a recombinant transdermal type VII collagen, said recombinant transdermal type VII collagen comprising: (a1) The amino acid sequence shown in SEQ ID NO.2; (a2) An amino acid sequence derived from SEQ ID NO.2 that has been substituted, deleted or added one or more amino acids and has type VII collagen activity; (a3) An amino acid sequence derived from SEQ ID NO.2 that has more than 90% sequence identity with the amino acid sequence shown in SEQ ID NO.2 and has type VII collagen activity.

[0007] Furthermore, the recombinant transdermal type VII collagen is also modified with a membrane-penetrating peptide, thereby further enhancing its transdermal function. The membrane-penetrating peptide can be modified at the carboxyl terminus (C-terminus) or amino terminus (N-terminus) of the above-mentioned recombinant transdermal type VII collagen, preferably at the amino terminus.

[0008] The membrane-penetrating peptide can be the membrane-penetrating peptide SPACE (amino acid sequence as shown in SEQ ID NO.5).

[0009] Therefore, the amino acid sequence of the recombinant transdermal type VII collagen is shown in SEQ ID NO.3.

[0010] In a second aspect, the present invention provides a nucleic acid molecule capable of encoding the aforementioned recombinant transdermal type VII collagen; specifically, the nucleotide sequence of the nucleic acid molecule is shown in SEQ ID NO.4.

[0011] A third aspect of the present invention provides a recombinant expression vector comprising the nucleic acid molecule.

[0012] A fourth aspect of the present invention provides a host cell containing the recombinant expression vector or having the nucleic acid molecule integrated into a chromosome, or being able to express the recombinant transdermal type VII collagen.

[0013] A fifth aspect of the present invention provides a method for preparing the recombinant transdermal type VII collagen, comprising: culturing the host cells to express the recombinant transdermal type VII collagen; and isolating and purifying the recombinant transdermal type VII collagen.

[0014] A sixth aspect of the present invention provides the use of the above-mentioned recombinant transdermal type VII collagen in the preparation of health foods, daily chemical products, medical aesthetic products or medical devices.

[0015] The beneficial technical effects of one or more of the above technical solutions are as follows: The above technical solution provides a recombinant transdermal type VII collagen, preferably containing a 54-amino acid sequence, tandemly with 6 copies, and with a membrane-penetrating peptide introduced at the N-terminus. Experimental verification shows that it has better transdermal performance and cell proliferation promotion performance compared to commercially available recombinant type VII collagen. The better transdermal properties allow it to enter the epidermis to exert its function and promote epidermal wound healing. Furthermore, the preparation method of the recombinant transdermal type VII collagen in the above technical solution is simple and easy to implement, making it very suitable for large-scale industrial production, thus possessing significant practical application value. Attached Figure Description

[0016] The accompanying drawings, which form part of this invention, are used to provide a further understanding of the invention. The illustrative embodiments of the invention and their descriptions are used to explain the invention and do not constitute an improper limitation of the invention.

[0017] Figure 1 Map of yeast expression plasmid pPIC9K-SPACE-324.

[0018] Figure 2 M: 10-180 kDa protein marker; Lanes 1-4: GS115 / pPIC9K-SPACE-324 fermentation broth supernatant after 24, 48, 72, and 96 hours of fermentation. Detailed Implementation

[0019] It should be noted that the following detailed description is illustrative and intended to provide further explanation of the invention. Unless otherwise specified, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention pertains.

[0020] It should be noted that the terminology used herein is for the purpose of describing particular embodiments only and is not intended to limit the exemplary embodiments according to this application. As used herein, the singular form is intended to include the plural form as well, unless the context clearly indicates otherwise. Furthermore, it should be understood that when the terms "comprising" and / or "including" are used in this specification, they indicate the presence of features, steps, operations, devices, components, and / or combinations thereof.

[0021] In a typical embodiment of the present invention, a recombinant transdermal type VII collagen is provided, the recombinant transdermal type VII collagen comprising: (a1) The amino acid sequence shown in SEQ ID NO.2; (a2) An amino acid sequence derived from SEQ ID NO.2 that has been substituted, deleted or added one or more amino acids and has type VII collagen activity; (a3) An amino acid sequence derived from SEQ ID NO.2 that has more than 90% sequence identity with the amino acid sequence shown in SEQ ID NO.2 and has type VII collagen activity.

[0022] The proteins shown in (a1)–(a3) above can be synthesized artificially, or their encoding genes can be synthesized first and then expressed biologically.

[0023] In this invention, amino acid substitution refers to the replacement of an amino acid at a certain position in an amino acid sequence with another amino acid, as long as the altered sequence completely or partially retains the activity of the original amino acid sequence. Amino acid substitution can be conserved amino acid substitution, meaning that compared to the original amino acid sequence, several amino acids are replaced by amino acids with similar or related properties to form a peptide.

[0024] In this invention, amino acid deletion may refer to the deletion of 1, 2 or 3 or more amino acids from the amino acid sequence, as long as the altered sequence completely or partially retains the activity of the original amino acid sequence.

[0025] In this invention, amino acid addition can refer to adding 1, 2 or 3 or more amino acids at any position at the C-terminus, N-terminus or between the C-terminus and N-terminus of the amino acid sequence, as long as the modified sequence completely or partially retains the activity of the original amino acid sequence.

[0026] Furthermore, the recombinant transdermal type VII collagen is also modified with a membrane-penetrating peptide, thereby further enhancing its transdermal function. The membrane-penetrating peptide can be modified at the carboxyl terminus (C-terminus) or amino terminus (N-terminus) of the above-mentioned recombinant transdermal type VII collagen, preferably at the amino terminus.

[0027] The membrane-penetrating peptide can be the membrane-penetrating peptide SPACE (amino acid sequence as shown in 6).

[0028] Therefore, in one specific embodiment of the present invention, the amino acid sequence of the recombinant transdermal type VII collagen is as shown in SEQ ID NO.3.

[0029] In another specific embodiment of the present invention, a nucleic acid molecule is provided, which is capable of encoding the above-mentioned recombinant transdermal type VII collagen; specifically, the nucleotide sequence of the nucleic acid molecule is shown in SEQ ID NO.4.

[0030] The nucleic acid molecule can be DNA, such as cDNA, genomic DNA, or recombinant DNA, etc., without specific limitations.

[0031] In another specific embodiment of the present invention, a recombinant expression vector is provided, wherein the recombinant expression vector comprises the nucleic acid molecule.

[0032] According to the present invention, the recombinant expression vector is obtained by effectively linking the aforementioned nucleic acid molecules to an expression vector. The expression vector is any one or more of a viral vector, plasmid, bacteriophage, kinase, or artificial chromosome. The viral vector may include adenovirus vector, retrovirus vector, or adeno-associated virus vector. The artificial chromosome includes bacterial artificial chromosomes, bacteriophage P1-derived vectors, yeast artificial chromosomes, or mammalian artificial chromosomes. In another specific embodiment of the present invention, the expression vector is a plasmid, specifically any plasmid vector from the pPIC series, and more specifically, the expression vector is the pPIC9K plasmid.

[0033] In another specific embodiment of the present invention, a host cell is provided, wherein the host cell contains the recombinant expression vector or the chromosome integrates the nucleic acid molecule or is capable of expressing the recombinant transdermal type VII collagen.

[0034] Furthermore, the host cell is a bacterial cell, a fungal cell, a mammalian cell, or an insect cell.

[0035] Furthermore, the fungal cells are yeast cells, specifically Pichia pastoris, such as Pichia pastoris X33, GS115, SMD1168, KM71, or KM71H. Pichia pastoris is one of the most common exogenous protein expression systems, possessing a clear expression regulation mechanism, low culture cost, high cell density, and significant protein expression potential. Simultaneously, Pichia pastoris also possesses a systematic gene editing system, a complete protein secretion expression mechanism, and the ability to modify exogenous proteins after translation, making it a widely used heterologous host in the food and pharmaceutical fields. Therefore, this invention preferentially uses Pichia pastoris.

[0036] In another specific embodiment of the present invention, a method for preparing the recombinant transdermal type VII collagen is provided, comprising: culturing the host cells to express the recombinant transdermal type VII collagen; and isolating and purifying the recombinant transdermal type VII collagen.

[0037] The separation and purification steps include removing impurities from proteins, centrifugation, hollow fiber membrane clarification, ultrafiltration membrane concentration, ion exchange chromatography, etc., which are not specifically limited here.

[0038] In another specific embodiment of the present invention, the above-mentioned recombinant transdermal type VII collagen is provided for use in the preparation of health foods, daily chemical products, medical aesthetic products or medical devices.

[0039] The aforementioned health food is a type of food that shares common characteristics with general foods. It can regulate the body's functions and is suitable for consumption by specific groups of people, but it is not intended to treat diseases.

[0040] The daily chemical products mentioned can be personal hygiene cleaners and cosmetics, specifically such as toothpaste, mouthwash, disinfectants, shampoos, hair lotions, hair gels, shower gels, soaps, facial cleansers, face masks, face creams, sunscreens, etc. In this invention, the cosmetics can be applied to the skin to achieve the effect of promoting cell proliferation; therefore, the cosmetics mentioned in this invention can be skin care products.

[0041] Meanwhile, in this invention, the cosmetic dosage form can be an aqueous solution, emulsion, ointment, etc. Furthermore, other cosmetic categories derived and prepared based on the above-mentioned basic cosmetic categories, such as facial cleanser, sunscreen, toner, face cream, eye cream, essence water (lotion), hand cream (ointment), body lotion, and face mask, are also within the scope of protection of this application.

[0042] In this invention, the medical aesthetic products refer to products used in the field of medical aesthetics, which are different from ordinary daily skincare products and beauty salon skincare products. Some commonly used medical aesthetic products include medical aesthetic masks and dressings. The use of these products requires professional medical personnel or relevant guidance. Medical aesthetic products, such as dressings, can be applied to those with impaired skin barrier function after procedures such as dermatitis, eczema, sensitive skin, and post-procedure treatments such as photofacial rejuvenation, laser beauty treatments, and chemical peels.

[0043] In this invention, the medical device can be a pharmaceutical device product; specifically, the pharmaceutical device product can be a topical dressing containing the aforementioned recombinant transdermal type VII collagen. This invention has demonstrated through experiments that this collagen can promote skin wound healing, has no cytotoxicity, and can be used to prepare medical dressings and other medical device products.

[0044] In another specific embodiment of the present invention, a medical device is provided, the medical device comprising the above-mentioned recombinant transdermal type VII collagen.

[0045] The present invention will be further described below with reference to embodiments. The present invention will be further illustrated by way of embodiments, but this does not limit the present invention to the scope of the embodiments described. Unless otherwise specified, the experimental methods used in the embodiments and comparative examples are conventional methods; the materials and reagents used, unless otherwise specified, are commercially available.

[0046] Example 1: Sequence design of recombinant transdermal type VII collagen The human type VII collagen sequence was analyzed to predict its active sites and functional regions. Based on the prediction, a GXY repeat sequence containing 54 amino acids was selected, as shown in SEQ ID NO.1. The amino acid sequence as shown in SEQ ID NO.2 was formed by repeating it tandemly 6 times. A SPACE transmembrane peptide was added to the N-terminus of the SEQ ID NO.2 sequence to form the amino acid sequence as shown in SEQ ID NO.3, thus obtaining the recombinant transdermal type VII collagen sequence.

[0047] Example 2: Construction, screening, and shake-flask fermentation of recombinant transdermal type VII collagen high-expression strains The recombinant humanized type VII collagen sequence obtained in Example 1 was codon-optimized according to the codon preference of Pichia pastoris. The optimized nucleotide sequence is shown in SEQ ID NO.4. The whole genome was synthesized by Nanjing Genscript Biotech Co., Ltd., and cloned downstream of the α signal peptide of the pPIC9K plasmid by homologous recombination to obtain the Pichia pastoris expression plasmid pPIC9K-SPACE-324. The vector map is shown below. Figure 1 As shown.

[0048] The expression vector pPIC9K-SPACE-324 was digested with salI to obtain the linearized fragment pPIC9K-SPACE-324-salI, which was then electroporated to... Pichia pastoris In GS115 competent cells, the cells were spread onto MD solid medium plates and incubated upside down at 30°C for 2-3 days. Transformants were observed to grow. The transformants were then transferred to solid medium plates containing 4 mg / mL genimycin G418 for screening. After incubation upside down at 30°C for 2-3 days, four transformants that grew well on the high-concentration antibiotic plates were selected and preserved.

[0049] The transformants were activated and inoculated into 10 mL of YPD liquid medium. Seed culture was carried out at 200 rpm and 30°C until OD reached [value missing]. 600= 8-10, inoculate 10% into 50 mL of BMGY (yeast extract 10 g / L, peptone 20 g / L, K2HPO4 3 g / L, KH2PO4 11.8 g / L, YNB 3.4 g / L, (NH4)2SO4 10 g / L, biotin 0.0004 g / L, glycerol 10 g / L) liquid medium, incubate at 200 rpm and 30℃ for 24 h, centrifuge at 4000 rpm for 2 min, discard the supernatant, and add 40 mL of BMGY (yeast extract 10 g / L, peptone 20 g / L, K2HPO4 3 g / L, KH2PO4 11.8 g / L, YNB 3.4 g / L, (NH4)2SO4 10 g / L, biotin 0.0004 g / L, methanol 10 g / L) The culture medium was prepared at 1 mL / L, and 1 mL of methanol inducer was added at 24, 48, 72, and 96 h, and samples were taken for SDS-PAGE analysis. Figure 2 Meanwhile, recombinant Pichia pastoris GS115 / pPIC9K-324 without SPACE-penetrating peptides was constructed according to the above preparation method, and the optimized nucleotide sequence of the recombinant humanized type VII collagen contained therein is shown in SEQ ID NO.6.

[0050] Example 3: High-density fermentation of recombinant Pichia pastoris GS115 / pPIC9K-324 and GS115 / pPIC9K-SPACE-324 in a 20L tank. Recombinant bacteria GS115 / pPIC9K-324 and GS115 / pPIC9K-SPACE-324 glycerol culture were streaked onto YPD solid agar plates for activation. Single colonies were picked and cultured in YPD liquid agar at 30°C and 200 rpm for primary seed culture. The bacterial OD... 600 When the bacterial cell OD reaches 5-10, it is inoculated again into YPD liquid medium at a 1% inoculum rate. The secondary seed culture is cultured at 30℃ and 200 rpm. 600 8-10 were introduced into a 20L tank for high-density culture.

[0051] Fermentation process and control: 10 L of BSM (85% H3PO4, 0.93 g / L calcium sulfate CaSO4, 18.2 g / L K2SO4, 14.9 g / L MgSO4·7H2O, 4.13 g / L KOH, 40 g / L glycerol) medium was added to a 20 L fermenter. The fermentation was carried out at 200 rpm, pH 5.0, 30℃, aeration rate of 0.5-2 vvm, and a pressure of 0.03 MPa. A 5% inoculum of secondary seed culture was added to initiate fermentation. Dissolved oxygen was controlled at 25% and the fermentation speed was linked to dissolved oxygen levels. When dissolved oxygen began to rebound, indicating complete depletion of the carbon source in the medium, continuous feeding (50% glycerol, containing 1.2% (V:V) PTM1) was initiated. When the wet weight of the bacteria reached approximately 200 g / L, glycerol feeding was stopped, and cultivation continued. Once the pH and dissolved oxygen levels recovered again, the fermentation speed was fixed at 900 rpm. At rpm, the dissolved oxygen was maintained within 25% by adjusting the feeding rate. Methanol (containing 1.2% (V:V) PTM1) was added at the beginning. The pH and temperature remained unchanged. After induction for 12 hours, the temperature was lowered to 25°C and induction continued for 72-96 hours. Fermentation was then stopped, and the fermentation supernatant was collected by centrifugation.

[0052] Example 4: Preparation of recombinant type VII collagen 324 and recombinant transdermal type VII collagen SPACE324 (1) Hollow fiber clarification: The fermentation supernatant obtained in Example 3 was filtered using a 250 kDa hollow fiber to remove residual bacteria and macromolecular impurities. (2) Ultrafiltration membrane concentration: The feed solution obtained in step (1) is concentrated using a 20 kDa ultrafiltration membrane pack to remove small molecule impurities; (3) Ion exchange chromatography: The feed solution obtained in step (2) is eluted using a cation exchange chromatography column at 40% and the elution peak is collected. (4) Ultrafiltration membrane desalination: The liquid collected in step (3) is desalinated by replacing it with pure water using a 20 kDa ultrafiltration membrane pack; The above-obtained liquid was freeze-dried to obtain collagen samples with a purity greater than 95%.

[0053] Example 5: Detection of the transdermal effect of recombinant transdermal type VII collagen The permeability of collagen samples was tested using piglet skin tissue. A sample with a concentration of 10 g / L was applied to the surface, with collagen without transmembrane peptides as a control. The accumulation of fluorescent markers in piglet skin was observed at 4 and 8 hours using a fluorescence microscope to evaluate the transdermal effect of the recombinant type VII humanized collagen provided by this invention.

[0054] The test results are shown in Table 1. As the test time increased, the mass of the fluorescent substances in both groups continued to increase. The increase in the experimental group was significantly greater than that in the control group, indicating that the recombinant type VII humanized collagen provided by this invention has a better transdermal absorption effect.

[0055] Table 1 Fluorescence Intensity

[0056] Example 6: Detection of Cell Proliferation Activity of Recombinant Transdermal Type VII Collagen The CCK-8 assay was used to detect the WST-8 compound, which was reduced by dehydrogenases in the mitochondria of live cells to produce a highly water-soluble orange formazan product. The intensity of the color of the product was directly proportional to the number of live cells. The proliferative activity of type VII humanized collagen was evaluated by measuring the absorbance at a wavelength of 450 nm using an ELISA reader.

[0057] Resuscitated human skin fibroblasts were cultured in DMEM medium containing 10% fetal bovine serum and incubated at 37°C until the logarithmic growth phase, then inoculated with 5 × 10⁻⁶ cells / mL. 3 Cells were seeded at a density of 100 μL / well in 96-well plates. Collagen samples were dissolved in phosphate buffer to a concentration of 10 mg / mL, and then added to the 96-well plates at progressively higher concentrations of 0.1, 0.25, 0.5, and 1.0 mg / mL. Wells without added collagen served as controls. Each concentration was treated in quadruplicates. After 48 h of incubation, 1 / 10 volume of CCK-8 solution was added to each well, and the plates were incubated at 37 °C for 2 h. The absorbance was measured at 450 nm using an enzyme-linked immunosorbent assay (ELISA) reader, and the results were recorded.

[0058] Table 2 shows the results of the effect of recombinant transdermal type VII collagen on the proliferation of human skin fibroblasts. After 48 hours of culture, under different concentrations of collagen, the cell viability of each group was significantly higher than that of the control group. Within the range of 0-0.5 mg / mL, the cell viability-promoting effect increased with increasing collagen concentration, and at 0.5 mg / mL, the cell growth rate was promoted by 15.9%. Therefore, this collagen can promote skin wound healing, has no cytotoxicity, and can be used to prepare medical dressings and other medical device products.

[0059] Table 2 Cell viability

[0060] The above description is merely a preferred embodiment of the present invention and is not intended to limit the invention. Various modifications and variations can be made to the present invention by those skilled in the art. Any modifications, equivalent substitutions, improvements, etc., made within the spirit and principles of the present invention should be included within the scope of protection of the present invention.

Claims

1. A recombinant transdermal type VII collagen, characterized in that, The recombinant transdermal type VII collagen includes: (a1) The amino acid sequence shown in SEQ ID NO.2; (a2) An amino acid sequence derived from SEQ ID NO.2 that has been substituted, deleted or added with one or more amino acids and has type VII collagen whitening activity; (a3) An amino acid sequence derived from SEQ ID NO.2 that has more than 90% sequence identity with the amino acid sequence shown in SEQ ID NO.2 and has type VII collagen whitening activity.

2. The recombinant transdermal type VII collagen as described in claim 1, characterized in that, The recombinant transdermal type VII collagen is further modified with a membrane-penetrating peptide. More specifically, the membrane-penetrating peptide is modified at the carboxyl terminus or amino terminus of the recombinant transdermal type VII collagen, preferably at the amino terminus.

3. The recombinant transdermal type VII collagen as described in claim 2, characterized in that, The membrane-penetrating peptide is SPACE. Furthermore, the amino acid sequence of the recombinant transdermal type VII collagen is shown in SEQ ID NO.

3.

4. A nucleic acid molecule, characterized in that, The nucleic acid molecule is capable of encoding the recombinant transdermal type VII collagen as described in any one of claims 1-3.

5. The nucleic acid molecule as described in claim 4, characterized in that, The nucleotide sequence of the nucleic acid molecule is shown in SEQ ID NO.

4.

6. A recombinant expression vector, characterized in that, The recombinant expression vector comprises the nucleic acid molecule as described in claim 4 or 5.

7. A host cell, characterized in that, The host cell contains the recombinant expression vector of claim 6 or a chromosome integrated with the nucleic acid molecule of claim 4 or 5, or is capable of expressing the recombinant transdermal type VII collagen of any one of claims 1-3.

8. A method for preparing the recombinant transdermal type VII collagen according to any one of claims 1-3, characterized in that, include: The host cells of claim 7 are cultured to express the recombinant transdermal type VII collagen; In addition, the recombinant transdermal type VII collagen was isolated and purified.

9. The use of the recombinant transdermal type VII collagen according to any one of claims 1-3 in the preparation of health foods, daily chemical products, medical aesthetic products or medical devices.

10. The application as described in claim 9, characterized in that, The daily chemical products are personal hygiene cleaners or cosmetics; further, the cosmetics are skin care products; The medical device is a pharmaceutical product; further, the medical device is a topical dressing containing the recombinant transdermal type VII collagen.

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