Specific sequence nucleic acid molecule and application thereof as PDRN

Abstract

The invention provides a specific sequence nucleic acid molecule and application thereof as a PDRN, and relates to the technical field of biology. A novel nucleic acid molecule with a specific sequence is obtained, and the sequence has certainty, biological activity and stable product quality. The PDRN of the specific sequence does not have cytotoxicity, is high in safety and has a wide application prospect. Moreover, compared with the extracted PDRN, the PDRN with a specific sequence is found for the first time, the product has a remarkable effect of promoting VEGF expression, cell damage can be effectively repaired, and a novel skin-care and repairing functional component is provided for the industries of cosmetics and medicines. The invention further provides a preparation method of the PDRN with the specific sequence, and the preparation method is simple and suitable for popularization, application and industrial production.

Description

[0001] This application is a divisional application of Chinese patent application filed on September 25, 2025, with application number "202511375770.9" and title "Specific Sequence Nucleic Acid Molecules and Their Application as PDRN". Technical Field

[0002] This application relates to the field of biotechnology, and in particular to specific sequence nucleic acid molecules and their applications as PDRNs. Background Technology

[0003] In recent years, PDRN has attracted widespread attention in the medical and cosmetic fields due to its unique biological activities. PDRN is usually derived from nucleic acid fragments from salmon sperm or mammalian tissues and has potential applications such as promoting tissue repair, anti-inflammation, and regenerative medicine. However, existing methods for preparing PDRN suffer from problems such as insufficient sequence specificity, unstable biological activity, or complex processes, which limit its clinical translation and commercial application.

[0004] Currently, traditional PDRN extraction methods mostly rely on physical shearing or enzymatic digestion, resulting in high product sequence randomness and uneven molecular weight distribution, which affects the reproducibility of its pharmacological effects. Furthermore, naturally derived PDRN may carry pathogen risks, and there are significant quality differences between different batches. Therefore, developing a PDRN synthesis method with a specific sequence is crucial to solving these problems. Summary of the Invention

[0005] The purpose of this invention is to provide new nucleic acid molecules with specific sequences that possess both good safety and repair functions.

[0006] On the one hand, this application provides a nucleic acid molecule comprising any one or more of the following sequences A1)-A11): A1) The nucleic acid molecule contains the nucleic acid sequence shown in SEQ ID NO.1 or a nucleic acid sequence that has 98% or more identity with SEQ ID NO.1; A2) The nucleic acid molecule contains the nucleic acid sequence shown in SEQ ID NO.2 or a nucleic acid sequence that has 98% or more identity with SEQ ID NO.2; A3) The nucleic acid molecule contains the nucleic acid sequence shown in SEQ ID NO.3 or a nucleic acid sequence that has 98% or more identity with SEQ ID NO.3; A4) The nucleic acid molecule contains the nucleic acid sequence shown in SEQ ID NO.4 or a nucleic acid sequence that has 98% or more identity with SEQ ID NO.4; A5) The nucleic acid molecule contains the nucleic acid sequence shown in SEQ ID NO. 5 or a nucleic acid sequence that has 98% or more identity with SEQ ID NO. 5; A6) The nucleic acid molecule contains the nucleic acid sequence shown in SEQ ID NO. 6 or a nucleic acid sequence that has 98% or more identity with SEQ ID NO. 6; A7) The nucleic acid molecule contains the nucleic acid sequence shown in SEQ ID NO.7 or a nucleic acid sequence that has 98% or more identity with SEQ ID NO.7; A8) The nucleic acid molecule contains the nucleic acid sequence shown in SEQ ID NO. 8 or a nucleic acid sequence that has 98% or more identity with SEQ ID NO. 8; A9) The nucleic acid molecule contains the nucleic acid sequence shown in SEQ ID NO. 9 or a nucleic acid sequence that has 98% or more identity with SEQ ID NO. 9; A10) A nucleic acid molecule that hybridizes under stringent conditions to any of the defined nucleic acid sequences in A1)-A9); Nucleic acid molecules that are complementary to any of the nucleic acid sequences defined in A11 and A1)-A9).

[0007] in, A1) The nucleic acid molecule contains the nucleic acid sequence shown in SEQ ID NO.1 or a nucleic acid sequence having 98%, 99%, 99.1%, 99.2%, 99.3%, 99.4%, 99.5%, 99.6%, 99.7%, 99.8% or 99.9% identity with SEQ ID NO.1; A2) The nucleic acid molecule contains the nucleic acid sequence shown in SEQ ID NO.2 or a nucleic acid sequence having 98%, 99%, 99.1%, 99.2%, 99.3%, 99.4%, 99.5%, 99.6%, 99.7%, 99.8% or 99.9% identity with SEQ ID NO.2; A3) The nucleic acid molecule contains the nucleic acid sequence shown in SEQ ID NO.3 or a nucleic acid sequence having 98%, 99%, 99.1%, 99.2%, 99.3%, 99.4%, 99.5%, 99.6%, 99.7%, 99.8% or 99.9% identity with SEQ ID NO.3; A4) The nucleic acid molecule contains the nucleic acid sequence shown in SEQ ID NO.4 or a nucleic acid sequence having 98%, 99%, 99.1%, 99.2%, 99.3%, 99.4%, 99.5%, 99.6%, 99.7%, 99.8% or 99.9% identity with SEQ ID NO.4; A5) The nucleic acid molecule contains the nucleic acid sequence shown in SEQ ID NO.5 or a nucleic acid sequence having 98%, 99%, 99.1%, 99.2%, 99.3%, 99.4%, 99.5%, 99.6%, 99.7%, 99.8% or 99.9% identity with SEQ ID NO.5; A6) The nucleic acid molecule contains the nucleic acid sequence shown in SEQ ID NO. 6 or a nucleic acid sequence having 98%, 99%, 99.1%, 99.2%, 99.3%, 99.4%, 99.5%, 99.6%, 99.7%, 99.8% or 99.9% identity with SEQ ID NO. 6; A7) The nucleic acid molecule contains the nucleic acid sequence shown in SEQ ID NO.7 or a nucleic acid sequence having 98%, 99%, 99.1%, 99.2%, 99.3%, 99.4%, 99.5%, 99.6%, 99.7%, 99.8% or 99.9% identity with SEQ ID NO.7; A8) The nucleic acid molecule contains the nucleic acid sequence shown in SEQ ID NO.8 or a nucleic acid sequence having 98%, 99%, 99.1%, 99.2%, 99.3%, 99.4%, 99.5%, 99.6%, 99.7%, 99.8% or 99.9% identity with SEQ ID NO.8; A9) The nucleic acid molecule contains the nucleic acid sequence shown in SEQ ID NO.9 or a nucleic acid sequence having 98%, 99%, 99.1%, 99.2%, 99.3%, 99.4%, 99.5%, 99.6%, 99.7%, 99.8% or 99.9% identity with SEQ ID NO.9; A10) A nucleic acid molecule that hybridizes under stringent conditions to any of the defined nucleic acid sequences in A1)-A9); Nucleic acid molecules that are complementary to any of the nucleic acid sequences defined in A11 and A1)-A9).

[0008] Identity refers to the degree of similarity between the nucleotide sequences of two nucleic acid molecules or the amino acid sequences of two protein molecules in molecular evolution studies.

[0009] Preferably, the nucleic acid molecule is a nucleic acid molecule that has the functions of promoting cell proliferation, promoting skin repair, promoting skin wound healing, improving skin inflammation, promoting angiogenesis and / or increasing the content of vascular endothelial growth factor.

[0010] More preferably, the nucleic acid molecule comprises a DNA molecule, specifically at least single-stranded DNA (ssDNA) or double-stranded DNA (dsDNA).

[0011] Those skilled in the art can prepare the nucleic acid molecules using conventional methods, including but not limited to artificial synthesis, amplification, gene expression, or mutation, without further limitation.

[0012] In a preferred embodiment, the method for preparing the nucleic acid molecule includes: amplification using salmon genomic DNA as a template.

[0013] Preferably, the salmon is chum salmon (Chum salmon). Oncorhynchus keta ).

[0014] Those skilled in the art can design amplification primers using conventional techniques, and no specific limitations are set here.

[0015] On the other hand, this application also provides a sodium salt of a nucleic acid molecule, wherein the nucleic acid molecule comprises at least one or more of the following sequences B1)-B4): B1) Contains any one or more nucleic acid sequences shown in SEQ ID NO.1, SEQ ID NO.2, SEQ ID NO.3, SEQ ID NO.4, SEQ ID NO.5, SEQ ID NO.6, SEQ ID NO.7, SEQ ID NO.8 or SEQ ID NO.9; B2) A nucleic acid sequence having 98% or more identity with the sequence described in B1); preferably, having 98%, 99%, 99.1%, 99.2%, 99.3%, 99.4%, 99.5%, 99.6%, 99.7%, 99.8%, or 99.9% identity. B3) Nucleic acid molecules that hybridize under stringent conditions with any of the specified nucleic acid sequences from B1) to B2); Nucleic acid molecules that are complementary to any of the defined nucleic acid sequences in B4 and B1-B2).

[0016] Those skilled in the art can prepare the sodium salt using conventional methods, and no further limitations are made herein.

[0017] On the other hand, this application also provides the use of nucleic acid molecules or their sodium salts as PDRN and / or in the preparation of PDRN-related products, wherein the nucleic acid molecules contain at least one or more of the following sequences B1)-B4): B1) Contains any one or more nucleic acid sequences shown in SEQ ID NO.1, SEQ ID NO.2, SEQ ID NO.3, SEQ ID NO.4, SEQ ID NO.5, SEQ ID NO.6, SEQ ID NO.7, SEQ ID NO.8 or SEQ ID NO.9; B2) A nucleic acid sequence having 98% or more identity with the sequence described in B1); preferably, having 98%, 99%, 99.1%, 99.2%, 99.3%, 99.4%, 99.5%, 99.6%, 99.7%, 99.8%, or 99.9% identity. B3) Nucleic acid molecules that hybridize under stringent conditions with any of the specified nucleic acid sequences from B1) to B2); Nucleic acid molecules that are complementary to any of the defined nucleic acid sequences in B4 and B1-B2).

[0018] On the other hand, this application also provides PDRN, which contains the nucleic acid molecule or the sodium salt.

[0019] Preferably, the PDRN is a PDRN that has the functions of promoting cell proliferation, promoting skin repair, promoting skin wound healing, improving skin inflammation, promoting angiogenesis and / or increasing the content of vascular endothelial growth factor.

[0020] On the other hand, this application also provides the use of the nucleic acid molecule, the sodium salt, the PDRN, or the product in the preparation of skin repair and / or skin inflammation improvement and / or skin wound healing products, wherein the nucleic acid molecule comprises at least one or more of the following sequences B1)-B4): B1) Contains any one or more nucleic acid sequences shown in SEQ ID NO.1, SEQ ID NO.2, SEQ ID NO.3, SEQ ID NO.4, SEQ ID NO.5, SEQ ID NO.6, SEQ ID NO.7, SEQ ID NO.8 or SEQ ID NO.9; B2) A nucleic acid sequence having 98% or more identity with the sequence described in B1); preferably, having 98%, 99%, 99.1%, 99.2%, 99.3%, 99.4%, 99.5%, 99.6%, 99.7%, 99.8%, or 99.9% identity. B3) Nucleic acid molecules that hybridize under stringent conditions with any of the specified nucleic acid sequences from B1) to B2); Nucleic acid molecules that are complementary to any of the defined nucleic acid sequences in B4 and B1-B2).

[0021] Furthermore, the applications include promoting cell proliferation and / or preparing products that promote cell proliferation.

[0022] Furthermore, the applications include promoting angiogenesis and / or preparing products that promote angiogenesis.

[0023] Furthermore, the application includes increasing vascular endothelial growth factor (VEGF) content and / or preparing products that increase VEGF content.

[0024] Furthermore, the products include cosmetics.

[0025] Furthermore, the products include pharmaceutical and medical device products.

[0026] The applications include the preparation of products that promote cell proliferation, skin repair, skin wound healing, improve skin inflammation, promote angiogenesis, and / or increase vascular endothelial growth factor.

[0027] Among these benefits, promoting skin repair, wound healing, improving skin inflammation, and / or promoting angiogenesis are achieved by increasing the content of vascular endothelial growth factor.

[0028] Preferably, increasing the vascular endothelial growth factor content includes increasing the expression level of vascular endothelial growth factor.

[0029] On the other hand, this application also provides the use of the nucleic acid molecule, the sodium salt, or the PDRN in promoting cell proliferation and / or preparing products that promote cell proliferation, wherein the nucleic acid molecule comprises at least one or more of the following sequences B1)-B4): B1) Contains any one or more nucleic acid sequences shown in SEQ ID NO.1, SEQ ID NO.2, SEQ ID NO.3, SEQ ID NO.4, SEQ ID NO.5, SEQ ID NO.6, SEQ ID NO.7, SEQ ID NO.8 or SEQ ID NO.9; B2) A nucleic acid sequence having 98% or more identity with the sequence described in B1); preferably, having 98%, 99%, 99.1%, 99.2%, 99.3%, 99.4%, 99.5%, 99.6%, 99.7%, 99.8%, or 99.9% identity. B3) Nucleic acid molecules that hybridize under stringent conditions with any of the specified nucleic acid sequences from B1) to B2); Nucleic acid molecules that are complementary to any of the defined nucleic acid sequences in B4 and B1-B2).

[0030] This application demonstrates that the nucleic acid molecule is non-toxic, highly safe, and has a relative cell proliferation rate of over 90%.

[0031] In a preferred embodiment, the cells are human immortalized keratinocytes (HaCat cells).

[0032] On the other hand, this application also provides the use of the nucleic acid molecule, the sodium salt, or the PDRN in skin repair and / or the preparation of skin repair products, wherein the nucleic acid molecule comprises at least one or more of the following sequences B1)-B4): B1) Contains any one or more nucleic acid sequences shown in SEQ ID NO.1, SEQ ID NO.2, SEQ ID NO.3, SEQ ID NO.4, SEQ ID NO.5, SEQ ID NO.6, SEQ ID NO.7, SEQ ID NO.8 or SEQ ID NO.9; B2) A nucleic acid sequence having 98% or more identity with the sequence described in B1); preferably, having 98%, 99%, 99.1%, 99.2%, 99.3%, 99.4%, 99.5%, 99.6%, 99.7%, 99.8%, or 99.9% identity. B3) Nucleic acid molecules that hybridize under stringent conditions with any of the specified nucleic acid sequences from B1) to B2); Nucleic acid molecules that are complementary to any of the defined nucleic acid sequences in B4 and B1-B2).

[0033] Preferably, the skin repair includes: increasing cytokine levels.

[0034] More preferably, the cytokines include vascular endothelial growth factor (VEGF).

[0035] In a preferred embodiment, the cells are human immortalized keratinocytes (HaCat cells).

[0036] On the other hand, this application also provides the use of the nucleic acid molecule, the sodium salt, or the PDRN in the preparation of skin wound healing products, wherein the nucleic acid molecule comprises at least one or more of the following sequences B1)-B4): B1) Contains any one or more nucleic acid sequences shown in SEQ ID NO.1, SEQ ID NO.2, SEQ ID NO.3, SEQ ID NO.4, SEQ ID NO.5, SEQ ID NO.6, SEQ ID NO.7, SEQ ID NO.8 or SEQ ID NO.9; B2) A nucleic acid sequence having 98% or more identity with the sequence described in B1); preferably, having 98%, 99%, 99.1%, 99.2%, 99.3%, 99.4%, 99.5%, 99.6%, 99.7%, 99.8%, or 99.9% identity. B3) Nucleic acid molecules that hybridize under stringent conditions with any of the specified nucleic acid sequences from B1) to B2); Nucleic acid molecules that are complementary to any of the defined nucleic acid sequences in B4 and B1-B2).

[0037] On the other hand, this application also provides the use of the nucleic acid molecule, the sodium salt, or the PDRN in the preparation of products for improving skin inflammation, wherein the nucleic acid molecule comprises at least one or more of the following sequences B1)-B4): B1) Contains any one or more nucleic acid sequences shown in SEQ ID NO.1, SEQ ID NO.2, SEQ ID NO.3, SEQ ID NO.4, SEQ ID NO.5, SEQ ID NO.6, SEQ ID NO.7, SEQ ID NO.8 or SEQ ID NO.9; B2) A nucleic acid sequence having 98% or more identity with the sequence described in B1); preferably, having 98%, 99%, 99.1%, 99.2%, 99.3%, 99.4%, 99.5%, 99.6%, 99.7%, 99.8%, or 99.9% identity. B3) Nucleic acid molecules that hybridize under stringent conditions with any of the specified nucleic acid sequences from B1) to B2); Nucleic acid molecules that are complementary to any of the defined nucleic acid sequences in B4 and B1-B2).

[0038] On the other hand, this application also provides the use of the nucleic acid molecule, the sodium salt, or the PDRN in the preparation of products that promote angiogenesis, wherein the nucleic acid molecule comprises at least one or more of the following sequences B1)-B4): B1) Contains any one or more nucleic acid sequences shown in SEQ ID NO.1, SEQ ID NO.2, SEQ ID NO.3, SEQ ID NO.4, SEQ ID NO.5, SEQ ID NO.6, SEQ ID NO.7, SEQ ID NO.8 or SEQ ID NO.9; B2) A nucleic acid sequence having 98% or more identity with the sequence described in B1); preferably, having 98%, 99%, 99.1%, 99.2%, 99.3%, 99.4%, 99.5%, 99.6%, 99.7%, 99.8%, or 99.9% identity. B3) Nucleic acid molecules that hybridize under stringent conditions with any of the specified nucleic acid sequences from B1) to B2); Nucleic acid molecules that are complementary to any of the defined nucleic acid sequences in B4 and B1-B2).

[0039] On the other hand, this application also provides the use of the nucleic acid molecule, the sodium salt, or the PDRN in increasing the content of vascular endothelial growth factor and / or in preparing products that increase the content of vascular endothelial growth factor, wherein the nucleic acid molecule comprises at least one or more of the following sequences B1)-B4): B1) Contains any one or more nucleic acid sequences shown in SEQ ID NO.1, SEQ ID NO.2, SEQ ID NO.3, SEQ ID NO.4, SEQ ID NO.5, SEQ ID NO.6, SEQ ID NO.7, SEQ ID NO.8 or SEQ ID NO.9; B2) A nucleic acid sequence having 98% or more identity with the sequence described in B1); preferably, having 98%, 99%, 99.1%, 99.2%, 99.3%, 99.4%, 99.5%, 99.6%, 99.7%, 99.8%, or 99.9% identity. B3) Nucleic acid molecules that hybridize under stringent conditions with any of the specified nucleic acid sequences from B1) to B2); Nucleic acid molecules that are complementary to any of the defined nucleic acid sequences in B4 and B1-B2).

[0040] Preferably, the increase in vascular endothelial growth factor (VEGF) content is achieved by promoting VEGF expression.

[0041] The nucleic acid molecules described in this application have a more significant effect on increasing the content of vascular endothelial growth factor compared to commercially available salmon PDRN products.

[0042] Preferably, the content of vascular endothelial growth factor is increased by more than or equal to 20%.

[0043] In a preferred embodiment, the cells are human immortalized keratinocytes (HaCat cells).

[0044] On the other hand, this application also provides a product comprising the nucleic acid molecule or the sodium salt or the PDRN.

[0045] Furthermore, the product also includes physiologically acceptable excipients.

[0046] Specifically, the excipients may be suitable solvents, propellants, solubilizers, co-solvents, emulsifiers, colorants, binders, disintegrants, fillers, lubricants, wetting agents, osmotic pressure regulators, stabilizers, flow aids, flavoring agents, preservatives, suspending agents, coating materials, fragrances, anti-adhesion agents, binding agents, penetration promoters, pH adjusters, buffers, plasticizers, surfactants, foaming agents, defoamers, thickeners, encapsulating agents, humectants, absorbents, diluents, flocculants and anti-flocculation agents, filter aids, release inhibitors, etc.

[0047] The product of this application can be prepared using general methods, wherein one or more diluents or carriers may be added. The product of this application can be prepared into any formulation commonly prepared in this field. For example, it can be formulated into creams, lotions, emulsions, masks, foundations, medical devices, hair cosmetics, etc. Specifically, skin lotions, skin softeners, hyaluronic acid injections, skin toners, astringents, lotions, moisturizing lotions, nourishing lotions, massage creams, nourishing creams, moisturizing creams, hand creams, foundations, serums, nourishing essences, masks, soaps, cleansing foams, cleansing milks, cleansing creams, lotions, or shower gels.

[0048] The products of this application may also be mixed with other ingredients commonly formulated in cosmetics, as needed. These may include, for example, oils, moisturizers, surfactants, organic pigments, inorganic pigments, UV absorbers, preservatives, bactericides, antioxidants, plant extracts, pH adjusters, alcohols, colorants, fragrances, blood circulation enhancers, cooling agents, antiperspirants, or may be mixed with purified water.

[0049] On the other hand, this application also provides the application of the product in promoting cell proliferation, promoting skin repair, promoting skin wound healing, improving skin inflammation, promoting angiogenesis and / or increasing the content of vascular endothelial growth factor.

[0050] Those skilled in the art can select the concentration of the nucleic acid molecule, the sodium salt, the PDRN, or the product according to the actual situation, and no specific limitation is made thereto.

[0051] In a preferred embodiment, the concentration of the nucleic acid molecule, the sodium salt, the PDRN, or the product may be 0.0001%-20%, preferably 0.0001%-5%. The concentration of the nucleic acid molecule, the sodium salt, the PDRN, or the product may be 0.0001%, 0.0002%, 0.0003%, 0.0004%, 0.0005%, 0.0006%, 0.0007%, 0.0008%, 0.0009%, 0.001%, 0.002%, 0.003%, 0.004%, 0.005%, 0.006%, 0.007%, 0.008%, 0.009%, 0.01%, etc. 0.02%, 0.03%, 0.04%, 0.05%, 0.06%, 0.07%, 0.08%, 0.09%, 0.1%, 0.2%, 0.3%, 0.4%, 0.5%, 0.6%, 0.7%, 0.8%, 0.9%, 1%, 2%, 3%, 4%, 5%, 6%, 7%, 8%, 9%, 10%, 11%, 12%, 13%, 14%, 15%, 16%, 17%, 18%, 19%, 20%, and any of these values.

[0052] The present invention has the following beneficial effects: This application yields a novel specific PDRN sequence with deterministic sequence characteristics and stable biological activity and product quality. This specific PDRN sequence exhibits no cytotoxicity, high safety, and broad application prospects. Furthermore, this application is the first to discover that this specific PDRN sequence promotes VEGF expression, effectively repairing cell damage and providing the cosmetics and pharmaceutical industries with a novel skincare and repair ingredient.

[0053] This application also provides a method for preparing PDRN with a specific sequence. The preparation method is simple and suitable for widespread application and industrial production. Attached Figure Description

[0054] The accompanying drawings, which are included to provide a further understanding of this application and form part of this application, illustrate exemplary embodiments and are used to explain this application, but do not constitute an undue limitation of this application. In the drawings: Figure 1 Electrophoresis diagram of PDRN for a specific sequence; Figure 2 The graph shows the relative VEGF production rate. NT is the blank control, 1-9 are SEQ ID No. 1-9 respectively, and PDRN is salmon PDRN. Detailed Implementation

[0055] To more clearly illustrate the overall concept of this application, a detailed description is provided below with reference to the accompanying drawings and embodiments. Numerous specific details are set forth in the following description to provide a more thorough understanding of the invention. However, it will be apparent to those skilled in the art that the invention can be practiced without one or more of these details. In other instances, certain technical features well-known in the art have not been described to avoid confusion with the invention.

[0056] It should be noted that the following detailed descriptions are illustrative and intended to provide further explanation of this application. 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 application pertains. Before further describing specific embodiments of the present invention, it should be understood that the scope of protection of the present invention is not limited to the specific embodiments described below; it should also be understood that the terminology used in the embodiments of the present invention is for describing specific embodiments and not for limiting the scope of protection of the present invention.

[0057] Unless otherwise specified, all reagents or instruments used in the following embodiments, unless otherwise indicated by the manufacturer, are commercially available products. Where specific conditions are not specified in the embodiments, they are performed under standard conditions or conditions recommended by the manufacturer.

[0058] The plasmids, restriction enzymes, PCR enzymes, column DNA extraction kits, and DNA gel recovery kits used in the following examples are commercial products. The specific operations were performed according to the kit instructions. Unless otherwise stated, the experimental methods, detection methods, and preparation methods disclosed in this invention all employ conventional techniques in molecular biology, biochemistry, chromatin structure and analysis, analytical chemistry, cell culture, recombinant DNA technology, and related fields. Specifically, they can be performed according to Molecular Cloning: A Laboratory Manual (Fourth Edition).

[0059] The term "water" as used in this invention includes any feasible type of water that can be used in the art, such as deionized water, distilled water, ion-exchange water, double-distilled water, high-purity water, and purified water.

[0060] The term "and / or" as used in this invention refers to and covers any and all possible combinations of one or more of the associated listed items, as well as the absence of a combination when interpreted in an alternative manner ("or").

[0061] In this invention, the terms "nucleic acid," "nucleic acid molecule," "nucleotide," "nucleotide sequence," and "polynucleotide" are used interchangeably and refer to the phosphate ester polymeric form of ribonucleosides (adenosine, guanosine, uridine, or cytidine; "RNA molecule") or deoxyribonucleosides (deoxyadenosine, deoxyguanosine, deoxythymidine, or deoxycytidine; "DNA molecule"), or any phosphate ester analogue thereof, such as single-stranded or double-stranded helical thiophosphates and thioesters. A single-stranded nucleic acid sequence refers to single-stranded DNA (ssDNA) or single-stranded RNA (ssRNA). Double-stranded DNA-DNA, DNA-RNA, and RNA-RNA helices are possible. The term nucleic acid molecule, particularly DNA or RNA molecule, refers only to the primary and secondary structures of the molecule and is not limited to any particular tertiary form. A "recombinant DNA molecule" is a DNA molecule that has undergone molecular biological manipulation. DNA includes, but is not limited to, cDNA, genomic DNA, plasmid DNA, synthetic DNA, and semi-synthetic DNA. The "product" of this disclosure comprises one or more nucleic acids as described herein.

[0062] Polydeoxyribonucleotides are single or double-stranded polymers consisting of phosphate, deoxyribose, and four bases (adenine, guanine, thymine, and cytosine).

[0063] Polyribonucleotides are single-chain polymers consisting of phosphate, ribose, and four bases (including adenine, guanine, cytosine, and uracil).

[0064] In this article, PDRN contains polydeoxyribonucleotides and / or polyribonucleotides and their associated salts.

[0065] Furthermore, in this paper, PDRN contains polydeoxyribonucleotides and / or polyribonucleotides and their associated sodium salts with a molecular weight of less than or equal to 1500 kDa.

[0066] Furthermore, those skilled in the art will recognize that PDRN often exists as a water-soluble salt formed with sodium ions, and the PDRN prepared by this invention also covers the case of sodium salts of nucleic acid molecules.

[0067] As is known in the art, the term "identity" refers to the relationship between two or more polypeptide sequences or two or more polynucleotide sequences, as determined by sequence comparison. In the art, "identity" also refers to the degree of sequence correlation between polypeptide or polynucleotide sequences, as determined by matching strings of such sequences. "Identity" can be readily calculated using known methods, including but not limited to those described in: *Computational Molecular Biology* (edited by Lesk, AM), Oxford University Press, New York (1988); *Biocomputing: Informatics and Genome Projects* (edited by Smith, DW), Academic Press, New York (1993); *Computer Analysis of Sequence Data, Part I* (edited by Griffin, AM and Griffin, HG), Humana Press, New Jersey (1994); *Sequence Analysis in Molecular Biology* (edited by von Heinje, G.), Academic Press (1987); and *Sequence Analysis Primer* (edited by Gribskov, M. and Devereux, J.), Stockton Press, New York (1991). Preferred methods for determining identity are designed to yield the best match between the tested sequences. Methods for determining identity have been incorporated into publicly available computer programs. Sequence alignment and percentage identity calculations can be performed using sequence analysis software such as the Megalign program of the LASERGENE Bioinformatics Computing Suite (DNASTAR, Madison, WI), the GCG program suite (Wisconsin Package version 9.0, Genetics Computer Group (GCG), Madison, WI), BLASTP, BLASTN, BLASTX (Altschul et al., J. Mol. Biol. 215:403 (1990)), and DNASTAR (DNASTAR, 1228 S. Park St. Madison, WI 53715 USA). In the context of this application, it will be understood that, when using sequence analysis software for analysis, unless otherwise stated, the results of the analysis will be based on the “default values” of the cited program. As used herein, “default values” refers to any set of values ​​or parameters initially loaded with the software upon initial initialization.

[0068] As used herein, the term "physiologically acceptable" means a molecular entity and composition that is physiologically tolerable and, when administered to humans, generally does not produce toxicity or sensitization or similar adverse reactions (such as stomach upset, dizziness, etc.). Optionally, as used herein, the term "physiologically acceptable" means approved by a government regulatory agency or approved in a pharmacopoeia or other generally recognized manner for use in animals or humans.

[0069] In the following embodiments, unless otherwise specified, % means wt%, i.e., weight percentage.

[0070] Example 1 Sequence Screening In this embodiment, salmon (a type of salmon) was downloaded from the public database NCBI (National Center of Biotechnology Information). Oncorhynchus keta The genome assembly information (version number: GCF_023373465.1) was used. Based on the genome annotation file information, annotation entries tagged with "gene" were first screened. Then, the chromosome information, coding start and end positions, and positive / negative strand information of the "gene" were analyzed through the annotation entries. Based on the "gene" location information and the genome sequence file, the DNA sequence corresponding to each "gene" was extracted, resulting in 663,364 gene sequences. The sequence length of each "gene" sequence was further calculated, and sequences ≤1500 bp in length were screened, yielding 13,046 candidate sequences. Further screening of these 13,046 candidate sequences resulted in 16 candidate sequences. Finally, sequence analysis was performed on these 16 candidate sequences, removing sequences with highly repetitive regions at both ends or within the sequence, resulting in 9 target sequences, as shown in SEQ ID No. 1-9.

[0071] Example 2: Preparation of Specific Sequences PCR amplification primers were designed based on the SEQ ID No. 1-9 sequences obtained in Example 1. The primer sequences are shown in Table 1.

[0072] Using salmon (Chum salmon) PDRN as a template (from Haiya Toning Co., Ltd., batch number APHL1252A), the nine selected sequences were amplified using 2 × Phanta Flash Master Mix (purchased from Nanjing Novizan Biotechnology Co., Ltd., catalog number P510-01). The amplification system and amplification program are shown in Tables 2 and 3 below.

[0073] Table 1

[0074] Table 2 Amplification System

[0075] Table 3 Amplification Procedure

[0076] Based on the above amplification system and procedure, the corresponding specific sequence PDRN was successfully prepared, and the results are as follows: Figure 1 As shown. Figure 1 The middle lane, from right to left, consists of: Marker (GeneRuler 1kb plus DNA Ladder, purchased from Thermo Fisher Scientific, catalog number SM1331), SEQ ID No. 1, SEQ ID No. 2, SEQ ID No. 3, SEQ ID No. 4, SEQ ID No. 5, SEQ ID No. 6, SEQ ID No. 7, SEQ ID No. 8, and SEQ ID No. 9.

[0077] After purification using a kit (purchased from Nanjing Novizan Biotechnology Co., Ltd., catalog number DC301-01), the PCR product was verified by Sanger sequencing by Suzhou Genewiz Biotechnology Co., Ltd. The results showed that the amplified sequence was consistent with the theoretical sequence.

[0078] Example 3 Cytotoxicity Evaluation Cytotoxicity safety was tested using the specific PDRN sequences SEQ ID No. 1-9 obtained in Example 2 and salmon PDRN (from Haiya Toning Co., Ltd., batch number APHL1252A) at a sample concentration of 0.01%.

[0079] Specifically, HaCat cells were used as the experimental subject, and the culture system was DMEM medium (with 10% fetal bovine serum added) under the conditions of 37°C and 5% CO2 in a carbon dioxide incubator. After 24 h of cell culture, the original medium was discarded and serum-free (DMEM) medium mixed with the sample was added, and the cells were cultured for another 24 h. The blank control was replaced with an equal volume of serum-free medium. The relative cell proliferation rate was detected using the CCK-8 assay (using the CCK-8 CellCounting Kit from Nanjing Novizan Biotechnology Co., Ltd., catalog number A311-01). The relative proliferation rate data are shown in Table 4.

[0080] Table 4 Results of Cytotoxicity Safety Tests

[0081] The relative cell proliferation rate data in Table 4 show that after treatment with the prepared specific sequence PDRN, the relative cell proliferation rate was above 90%, indicating that the above samples were not cytotoxic and had high safety.

[0082] Example 4: Efficacy Evaluation The efficacy was tested using PDRN samples with specific sequences obtained in Example 2 (SEQ ID No. 1-9) and salmon PDRN samples (from Haiya Tou Ning Co., Ltd., batch number APHL1252A) at a sample concentration of 0.01%.

[0083] Specifically, HaCat cells were used as the experimental subject, and the culture system was DMEM medium (with 10% fetal bovine serum added). The culture conditions were a CO2 incubator at 37°C and 5% CO2. The sample processing procedure was as follows: after 24 h of cell culture, the original culture medium was discarded, and serum-free (DMEM) medium containing the sample was added for another 24 h of culture. An equal volume of serum-free medium was added to the blank control. After the incubation period, the cell supernatant was collected, and the VEGF content was determined using a human VEGF ELISA kit according to the instructions. The test data are shown in Table 5. A graph was plotted from the data in Table 5. Figure 2 .

[0084] Table 5 Results of the test on the efficacy of promoting VEGF repair

[0085] Note: * indicates p<0.05 compared with the blank control group; ** indicates p<0.01 compared with the blank control group; # indicates p<0.05 compared with the salmon PDNR group; ## indicates p<0.01 compared with the salmon PDRN group; ### indicates p<0.001 compared with the salmon PDRN group.

[0086] The active expression of cytokines in wound repair makes them a new reference parameter for inferring damage levels, serving as a molecular-level "living response" indicator in in vivo injuries. VEGF is a key factor in the late inflammatory and proliferative phases of wound healing, playing a crucial role in granulation tissue formation, wound healing, angiogenesis, collagen fiber synthesis, and the repair of hair follicles and skin appendages. Therefore, detecting VEGF expression levels can effectively reflect the progress of cellular repair. The relative VEGF content data in Table 5 show that all nine PDRN samples with specific sequences prepared in Example 2 have the effect of promoting VEGF expression. Among them, the promoting effects of samples SEQ ID No. 1, SEQ ID No. 5, SEQ ID No. 6, SEQ ID No. 7, SEQ ID No. 8, and SEQ ID No. 9 are significant. Figure 2This indicates that the preparation of specific sequence samples has excellent effects on skin repair, skin wound healing, improvement of skin inflammation, and promotion of angiogenesis.

[0087] The above description is merely an embodiment of this application and is not intended to limit the scope of this application. Various modifications and variations can be made to this application by those skilled in the art. Any modifications, equivalent substitutions, improvements, etc., made within the spirit and principles of this application should be included within the scope of the claims of this application.

Claims

1. Use of a nucleic acid molecule or a sodium salt thereof in the manufacture of a PDRN- related product, characterized in that, The nucleic acid molecule comprises at least one or more of the following B1) - B4): B1) comprises any one or more of the nucleic acid sequences shown as SEQ ID NO. 2, SEQ ID NO. 3, SEQ ID NO. 4, SEQ ID NO. 5, SEQ ID NO. 6, SEQ ID NO. 7, SEQ ID NO. 8, or SEQ ID NO. 9; B2) a nucleic acid sequence that is 98% or more identical to the sequence of B1); B3) a nucleic acid molecule that hybridizes to any of the nucleic acid sequences defined in B1) - B2) under stringent conditions; B4) a nucleic acid molecule that is complementary to any of the nucleic acid sequences defined in B1) - B2).

2. Use of a nucleic acid molecule or a sodium salt thereof for the preparation of a skin repair and / or skin wound healing product, characterized in that, The nucleic acid molecule comprises at least one or more of the following B1) - B4): B1) comprises any one or more of the nucleic acid sequences shown as SEQ ID NO. 2, SEQ ID NO. 3, SEQ ID NO. 4, SEQ ID NO. 5, SEQ ID NO. 6, SEQ ID NO. 7, SEQ ID NO. 8, or SEQ ID NO. 9; B2) a nucleic acid sequence that is 98% or more identical to the sequence of B1); B3) a nucleic acid molecule that hybridizes to any of the nucleic acid sequences defined in B1) - B2) under stringent conditions; B4) a nucleic acid molecule that is complementary to any of the nucleic acid sequences defined in B1) - B2).

3. Use according to claim 2, characterized in that, The use also includes the preparation of a product that promotes cell proliferation.

4. Use according to claim 2, characterized in that, The use also includes the preparation of a product that promotes angiogenesis.

5. Use according to claim 2, characterized in that, The use also includes the preparation of a product that increases the content of vascular endothelial growth factor.

6. Use of a nucleic acid molecule or of its sodium salt for the preparation of a cosmetic and / or pharmaceutical product, characterized in that, The nucleic acid molecule comprises at least one or more of the following B1) - B4): B1) comprises any one or more of the nucleic acid sequences shown as SEQ ID NO. 2, SEQ ID NO. 3, SEQ ID NO. 4, SEQ ID NO. 5, SEQ ID NO. 6, SEQ ID NO. 7, SEQ ID NO. 8, or SEQ ID NO. 9; B2) a nucleic acid sequence that is 98% or more identical to the sequence of B1); B3) a nucleic acid molecule that hybridizes to any of the nucleic acid sequences defined in B1) - B2) under stringent conditions; B4) a nucleic acid molecule that is complementary to any of the nucleic acid sequences defined in B1) - B2).

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