Recombinant coral-spider silk fusion protein and functional alginate fiber and application thereof

Abstract

The invention relates to the technical field of biology, in particular to a recombinant coral-spider silk fusion protein and functional alginate fiber and application thereof. The recombinant coral-spider silk fusion protein provided by the invention comprises the recombinant coral-spider silk fusion protein formed by fusing spider silk protein or rattan silk protein and coral protein. The invention designs a fusion protein only containing spider silk protein and coral protein. The recombinant coral-spider silk fusion protein is applied to alginate fibers, colored spinning of the alginate fibers can be achieved, and the spun fibers are high in color fastness, superior to original fibers in strength and elongation and bright and stable in color.

Description

TECHNICAL FIELD

[0001] The present application relates to the field of biotechnology, in particular to a recombinant coral-spider silk fusion protein and functional seaweed fiber and uses thereof. BACKGROUND

[0002] Coral in the sea includes two states of medusa and hydrozoan, both of which have beautiful colors and form a beautiful landscape on the marine reef. Unlike other animals and plants that use small molecule pigments to color, the color of coral is determined by fluorescent proteins (FP) and chromoproteins (CP). These proteins have very high stability and structural homology.

[0003] The first fluorescent protein (FP) discovered was green fluorescent protein GFP isolated from Euplectella asperula (medusa also known as Victoria tube jellyfish) in 1995. This protein has been widely used in in vivo and in vitro live labeling and imaging techniques due to its single coloration and stable light emission properties. In addition, more than 30 fluorescent proteins from corals have been discovered and modified, and their fluorescent light includes red, orange, yellow, green, blue and other beautiful colors, which are also commonly used as biological markers. In addition to fluorescent proteins, chromoproteins (CP) are also common proteins in corals. They do not have the special effect of emitting fluorescence after being irradiated by a specific light, but they can present a specific color under visible light for a long time. These chromoproteins have brighter colors than fluorescent proteins and have higher light stability. Currently, there are fewer types of chromoproteins reported, which include red, orange, blue, purple and other different colors.

[0004] The monomer structure of fluorescent proteins and chromoproteins is very similar. They both have a β-barrel structure, which encloses a chromophore composed of three adjacent amino acids. The three chromophore amino acid residues cyclize to form an imidazolinone ring structure, which is then oxidized by molecular oxygen to form a mature chromophore, thereby bringing bright colors.

[0005] The chromoprotein and fluorescent protein of corals have bright color and charming fluorescence characteristics formed naturally, and their special light effects and aesthetic characteristics are also favored by fashion designers and artists, but how to make these proteins into textiles and artworks is a problem in the industry. The structure of the coral protein alone is not enough to form a fiber, film or other solid material structure, and needs to be compounded or fused with other materials to function. Seaweed fiber is a very suitable material: seaweed fiber is soft, skin-friendly and comfortable, and has natural antibacterial and flame-retardant properties. However, the biggest difficulty of seaweed fiber is that it is difficult to dye with conventional dyes, and the brightness and naturalness of color are difficult to meet the needs of textiles. Some researchers have tried to add natural colored proteins to seaweed fiber in order to produce seaweed fiber with bright and natural colors. However, it was found that the natural colored protein fused with seaweed fiber had low light stability and was prone to bleaching. SUMMARY

[0006] The present application provides a recombinant coral-spider silk fusion protein and functional seaweed fiber and uses thereof.

[0007] In a first aspect, the present application provides a recombinant coral-spider silk fusion protein, characterized in that it comprises a fusion of a spider silk protein or a recombinant spider silk protein and a coral protein to form a recombinant coral-spider silk fusion protein.

[0008] In some embodiments, the spider silk protein or spider silk-like protein comprises a repeat region of a spider silk protein or a moth silk protein.

[0009] In some embodiments, the N-terminus of the spider silk protein or spider silk-like protein further comprises a spider globin NT; and / or, the C-terminus of the spider silk protein or spider silk-like protein further comprises a spider globin CT.

[0010] In some embodiments, the spider globin is selected from the NT or CT of a natural spider silk; and / or, the repeat region of the spider silk protein is selected from the repeat region of any kind of spider silk protein, including but not limited to at least one of MaSp1, MaSp4, Flag, DmarAmSp, MiSp, AcSp, PySp or TuSp; the repeat region of the moth silk protein includes but is not limited to Ev1; Optionally, the amino acid sequence of the MaSp1 is as set forth in SEQ ID NO. 19 or a sequence having at least 85%, 88%, 90%, 93%, 95%, 96%, 97%, 98%, 99%, 99.9%, 99.99% or more, or 100% identity thereto; and the nucleotide sequence thereof is as set forth in SEQ ID NO. 20 or a sequence having at least 85%, 88%, 90%, 93%, 95%, 96%, 97%, 98%, 99%, 99.9%, 99.99% or more, or 100% identity thereto; Optionally, the amino acid sequence of the MaSp4 is as set forth in SEQ ID NO. 21 or a sequence having at least 85%, 88%, 90%, 93%, 95%, 96%, 97%, 98%, 99%, 99.9%, 99.99% or more, or 100% identity thereto; and the nucleotide sequence thereof is as set forth in SEQ ID NO. 22 or a sequence having at least 85%, 88%, 90%, 93%, 95%, 96%, 97%, 98%, 99%, 99.9%, 99.99% or more, or 100% identity thereto; Optionally, the amino acid sequence of the DmarAmSp is as set forth in SEQ ID NO. 23 or a sequence having at least 85%, 88%, 90%, 93%, 95%, 96%, 97%, 98%, 99%, 99.9%, 99.99% or more, or 100% identity thereto; and the nucleotide sequence thereof is as set forth in SEQ ID NO. 14 or a sequence having at least 85%, 88%, 90%, 93%, 95%, 96%, 97%, 98%, 99%, 99.9%, 99.99% or more, or 100% identity thereto; Optionally, the amino acid sequence of the Flag is as set forth in SEQ ID NO. 25 or a sequence having at least 85%, 88%, 90%, 93%, 95%, 96%, 97%, 98%, 99%, 99.9%, 99.99% or more, or 100% identity thereto; and the nucleotide sequence thereof is as set forth in SEQ ID NO. 26 or a sequence having at least 85%, 88%, 90%, 93%, 95%, 96%, 97%, 98%, 99%, 99.9%, 99.99% or more, or 100% identity thereto; Optionally, the amino acid sequence of the Evl is as set forth in SEQ ID NO. 27 or a sequence having at least 85%, 88%, 90%, 93%, 95%, 96%, 97%, 98%, 99%, 99.9%, 99.99% or more, or 100% identity thereto; and / or the nucleotide sequence of the Evl is as set forth in SEQ ID NO. 28 or a sequence having at least 85%, 88%, 90%, 93%, 95%, 96%, 97%, 98%, 99%, 99.9%, 99.99% or more, or 100% identity thereto. Optionally, the amino acid sequence of the globin NT is as set forth in SEQ ID NO. 29 or a sequence having at least 85%, 88%, 90%, 93%, 95%, 96%, 97%, 98%, 99%, 99.9%, 99.99% or more, or 100% identity thereto; and / or the nucleotide sequence of the globin NT is as set forth in SEQ ID NO. 30 or a sequence having at least 85%, 88%, 90%, 93%, 95%, 96%, 97%, 98%, 99%, 99.9%, 99.99% or more, or 100% identity thereto. Optionally, the amino acid sequence of the globin CT is as set forth in SEQ ID NO. 31 or a sequence having at least 85%, 88%, 90%, 93%, 95%, 96%, 97%, 98%, 99%, 99.9%, 99.99% or more, or 100% identity thereto; and / or the nucleotide sequence of the globin CT is as set forth in SEQ ID NO. 32 or a sequence having at least 85%, 88%, 90%, 93%, 95%, 96%, 97%, 98%, 99%, 99.9%, 99.99% or more, or 100% identity thereto.

[0011] In some embodiments, the coral protein is selected from a coral fluorescent protein and / or a coral chromoprotein. Optionally, the coral fluorescent protein includes, but is not limited to, at least one of coral fluorescent protein sfGFP, coral fluorescent protein eforRed, coral fluorescent protein amilCP, coral fluorescent protein mScarlet3, coral fluorescent protein FwYellow, coral fluorescent protein Yukon OFP, and coral fluorescent protein mTagBFP2. Optionally, the coral chromoprotein includes, but is not limited to, at least one of coral chromoprotein spisPink, coral chromoprotein gfasPurple, coral chromoprotein amilCP orange, and coral chromoprotein amilCP. Optionally, the amino acid sequence of the coral fluorescent protein sfGFP is as set forth in SEQ ID NO. 1 at positions 350-587 or a sequence having at least 85%, 88%, 90%, 93%, 95%, 96%, 97%, 98%, 99%, 99.9%, 99.99% or more, or 100% identity thereto; and the nucleotide sequence thereof is as set forth in SEQ ID NO. 2 at positions 1048-1761 or a sequence having at least 85%, 88%, 90%, 93%, 95%, 96%, 97%, 98%, 99%, 99.9%, 99.99% or more, or 100% identity thereto; Optionally, the amino acid sequence of the coral fluorescent protein eforRed is as set forth in SEQ ID NO. 5 at positions 227-453 or a sequence having at least 85%, 88%, 90%, 93%, 95%, 96%, 97%, 98%, 99%, 99.9%, 99.99% or more, or 100% identity thereto; and the nucleotide sequence thereof is as set forth in SEQ ID NO. 6 at positions 679-1359 or a sequence having at least 85%, 88%, 90%, 93%, 95%, 96%, 97%, 98%, 99%, 99.9%, 99.99% or more, or 100% identity thereto; Optionally, the amino acid sequence of the coral fluorescent protein amilCP is as set forth in SEQ ID NO. 7 at positions 9-229 or a sequence having at least 85%, 88%, 90%, 93%, 95%, 96%, 97%, 98%, 99%, 99.9%, 99.99% or more, or 100% identity thereto; and the nucleotide sequence thereof is as set forth in SEQ ID NO. 8 at positions 25-687 or a sequence having at least 85%, 88%, 90%, 93%, 95%, 96%, 97%, 98%, 99%, 99.9%, 99.99% or more, or 100% identity thereto; Optionally, the amino acid sequence of the coral fluorescent protein mScarlet3 is as set forth in SEQ ID NO. 9 at positions 317-545 or a sequence having at least 85%, 88%, 90%, 93%, 95%, 96%, 97%, 98%, 99%, 99.9%, 99.99% or more, or 100% identity thereto; and the nucleotide sequence thereof is as set forth in SEQ ID NO. 10 at positions 949-1635 or a sequence having at least 85%, 88%, 90%, 93%, 95%, 96%, 97%, 98%, 99%, 99.9%, 99.99% or more, or 100% identity thereto; Optionally, the coral fluorescent protein FwYellow is as set forth in SEQ ID NO. 13 from position 342 to 577 or a sequence having at least 85%, 88%, 90%, 93%, 95%, 96%, 97%, 98%, 99%, 99.9%, 99.99% or more, or 100% identity thereto; and the nucleotide sequence thereof is as set forth in SEQ ID NO. 14 from position 1024 to 1731 or a sequence having at least 85%, 88%, 90%, 93%, 95%, 96%, 97%, 98%, 99%, 99.9%, 99.99% or more, or 100% identity thereto; Optionally, the coral fluorescent protein Yukon OFP is as set forth in SEQ ID NO. 15 from position 369 to 595 or a sequence having at least 85%, 88%, 90%, 93%, 95%, 96%, 97%, 98%, 99%, 99.9%, 99.99% or more, or 100% identity thereto; and the nucleotide sequence thereof is as set forth in SEQ ID NO. 16 from position 1105 to 1785 or a sequence having at least 85%, 88%, 90%, 93%, 95%, 96%, 97%, 98%, 99%, 99.9%, 99.99% or more, or 100% identity thereto; Optionally, the coral fluorescent protein mTagBFP2 is as set forth in SEQ ID NO. 17 from position 400 to 636 or a sequence having at least 85%, 88%, 90%, 93%, 95%, 96%, 97%, 98%, 99%, 99.9%, 99.99% or more, or 100% identity thereto; and the nucleotide sequence thereof is as set forth in SEQ ID NO. 18 from position 1198 to 1908 or a sequence having at least 85%, 88%, 90%, 93%, 95%, 96%, 97%, 98%, 99%, 99.9%, 99.99% or more, or 100% identity thereto; Optionally, the coral color protein spisPink is as set forth in SEQ ID NO. 9 from position 1 to 224 or a sequence having at least 85%, 88%, 90%, 93%, 95%, 96%, 97%, 98%, 99%, 99.9%, 99.99% or more, or 100% identity thereto; and the nucleotide sequence thereof is as set forth in SEQ ID NO. 10 from position 1 to 672 or a sequence having at least 85%, 88%, 90%, 93%, 95%, 96%, 97%, 98%, 99%, 99.9%, 99.99% or more, or 100% identity thereto; Optionally, the coral fluorescent protein gfasPurple is as set forth in SEQ ID NO. 11 or a sequence having at least 85%, 88%, 90%, 93%, 95%, 96%, 97%, 98%, 99%, 99.9%, 99.99% or more, or 100% identity thereto; and the nucleotide sequence thereof is as set forth in SEQ ID NO. 12 or a sequence having at least 85%, 88%, 90%, 93%, 95%, 96%, 97%, 98%, 99%, 99.9%, 99.99% or more, or 100% identity thereto. Optionally, the coral fluorescent protein amilCP orange is as set forth in SEQ ID NO. 13 or a sequence having at least 85%, 88%, 90%, 93%, 95%, 96%, 97%, 98%, 99%, 99.9%, 99.99% or more, or 100% identity thereto; and the nucleotide sequence thereof is as set forth in SEQ ID NO. 14 or a sequence having at least 85%, 88%, 90%, 93%, 95%, 96%, 97%, 98%, 99%, 99.9%, 99.99% or more, or 100% identity thereto. Optionally, the coral fluorescent protein amilCP is as set forth in SEQ ID NO. 17 or a sequence having at least 85%, 88%, 90%, 93%, 95%, 96%, 97%, 98%, 99%, 99.9%, 99.99% or more, or 100% identity thereto; and the nucleotide sequence thereof is as set forth in SEQ ID NO. 18 or a sequence having at least 85%, 88%, 90%, 93%, 95%, 96%, 97%, 98%, 99%, 99.9%, 99.99% or more, or 100% identity thereto.

[0012] In some embodiments, the coral protein is fused to at least one end of the spider silk protein or spider silk-like protein.

[0013] In some embodiments, the recombinant coral-spider silk fusion protein has a structure from N-terminus to C-terminus selected from the group consisting of coral protein-NT globin-repeating region-CT globin, NT globin-repeating region-CT globin-coral protein, coral protein-repeating region-CT globin, NT globin-repeating region-coral protein, coral protein A-repeating region-coral protein B; The coral protein A and the coral protein B are respectively selected from the group consisting of a coral fluorescent protein and / or a coral chromoprotein; Preferably, the coral protein A and the coral protein B are the same or different.

[0014] In some embodiments, a linker peptide is further provided between the spider silk protein or spider silk-like protein and the spider venom protein. In some embodiments, at least one end of the recombinant coral-spider silk fusion protein is further fused with a His tag.

[0015] In some embodiments, the recombinant coral-spider silk fusion protein is selected from any one of NT-2Rep-CT-sfGFP, sfGFP-NT-2Rep-CT, NT-2Rep-eforRed, amilCP-2Rep-CT, spisPink-2Rep-mScarlet3, gfasPurple-D3r-mScarlet3, amilCP orange-Dmar-FwYellow, amilCP orange-Flag-YukonOFP, amilCP-EV1-mTagBFP2.

[0016] The embodiments of the present application provide a biomaterial, comprising any one of the following: 1) a nucleic acid molecule encoding the recombinant coral-spider silk fusion protein; optionally, the nucleic acid molecule is DNA or RNA; 2) an expression cassette, a recombinant vector, a recombinant microorganism or a transgenic cell line expressing the nucleic acid molecule encoding the recombinant coral-spider silk fusion protein; 3) an expression cassette, a recombinant vector, a recombinant microorganism or a transgenic cell line comprising the nucleic acid molecule of 1); 4) a recombinant vector, a recombinant microorganism or a transgenic cell line comprising the expression cassette of 2) or 3); 5) a host cell comprising the recombinant vector of 2), 3) or 4).

[0017] In some embodiments, the host cell of the recombinant microorganism is selected from the family Enterobacteriaceae Escherichia; Optionally, the host cell is selected from Escherichia coli; optionally, the Escherichia coli is Escherichia coli BL21 (DE3).

[0018] The embodiments of the present application provide a preparation method of the recombinant coral-spider silk fusion protein, comprising: fermenting a recombinant microorganism expressing the recombinant coral-spider silk fusion protein.

[0019] The embodiments of the present application provide a preparation method of a functional seaweed fiber, comprising: mixing the recombinant coral-spider silk fusion protein or the recombinant coral-spider silk fusion protein prepared by the preparation method with a polysaccharide compound to prepare a spinning solution; and preparing a polysaccharide fiber containing the recombinant coral-spider silk fusion protein by using the spinning solution.

[0020] In some embodiments, the mass fraction of the recombinant corona-spider silk fusion protein in the spinning solution is 0.15-1.5%, and the mass fraction of the polysaccharide compound is 4-10%. And / or, the polysaccharide compound is a polysaccharide compound extracted from seaweed; preferably, the polysaccharide compound extracted from seaweed is at least one selected from sodium alginate and trehalose.

[0021] In some embodiments, before the step of preparing the polysaccharide fiber containing the recombinant corona-spider silk fusion protein by using the spinning solution, the spinning solution is subjected to a defoaming step.

[0022] In some embodiments, the spinning solution is used to obtain the polysaccharide fiber containing the recombinant corona-spider silk fusion protein by wet spinning.

[0023] In some embodiments, the step of preparing the polysaccharide fiber containing the recombinant corona-spider silk fusion protein by using the spinning solution includes the following steps: adding the spinning solution into a spinning tube, wet spinning, and solidifying the spinning through a coagulation bath. A spinning needle with a pore size of 0.1-0.5 mm is used. The coagulation bath is a solution containing Ca 2+ , Cl - or PO 4- ions; preferably, the first coagulation bath is a 5-10wt% calcium chloride solution, and the second coagulation bath is a 0-10wt% calcium chloride solution. In the spinning, the spinning speed is 10-40 m / min. In the spinning, the draw ratio is 1:1.1-1.5.

[0024] The functional seaweed fiber prepared by the preparation method of the functional seaweed fiber is provided in the embodiments of the present application.

[0025] The functional seaweed fiber has any one of the following uses: (1) in textile use; (2) in art use; (3) in hand-made toy use; (4) in vehicle decoration use; (5) in household sofa use; (6) in safety warning product use.

[0026] The technical scheme of the present application has the following advantages: 1. The present application provides a recombinant coral-spider silk fusion protein, comprising fusion of spider silk protein or recombinant spider silk protein and coral protein to form a recombinant coral-spider silk fusion protein. The present application designs a fusion protein containing only spider silk protein and coral protein. The recombinant coral-spider silk fusion protein is applied to seaweed fibers, which can realize colored spinning of seaweed fibers, and the color fastness of the spun fibers is high, the strength and elongation are better than those of the original fibers, and the color is bright and stable.

[0027] Further, the coral protein can be directly fused to the N or C terminal of the complete spider silk protein (containing NT globin-repeating region-CT globin), to form a recombinant protein of "coral protein-NT globin-repeating region-CT globin" or "NT globin-repeating region-CT globin-coral protein", or can replace one end of the globin to form "coral protein-repeating region-CT globin" or "NT globin-repeating region-coral protein". These proteins can be expressed in water-soluble form in an E. coli chassis, and realize colored spinning of seaweed fibers. The color fastness of the spun fibers is high, the toughness is better than that of the original fibers, and the color is bright and stable.

[0028] More preferably, the NT terminal globin and CT terminal globin of the spider silk protein are completely replaced to form a fusion protein of "coral protein 1-repeating region-coral protein 2", which is found to have various properties similar to the original spider silk protein. Through various color combinations, including but not limited to combinations of 5 kinds of fluorescent proteins and 4 kinds of color proteins, colored spinning of seaweed fibers is realized, and green, blue, orange, yellow, purple and red fibers of various colors are obtained. Through the mutual combination of various proteins, the types of protein colors far exceed the natural coral protein colors, the color fastness of the spun fibers is high, the toughness is better than that of the original fibers, and the color is bright and stable.

[0029] 2. The present application provides a method for preparing functional seaweed fibers. The coral protein fused spider silk protein can be dissolved in the same spinning solution as alginic acid and coagulated in the same coagulation bath, which also shows that the recombinant coral-spider silk fusion protein has the properties of spider silk protein. The spinning solution of spider silk is weakly alkaline with a pH of 8-10, while the spinning solution of alginic acid has a similar pH environment; the pH of the spider silk when ejected from the spinner is 3-5, and certain ionic components are needed to assist spinning. The addition of Ca 2+ , Cl - , PO 4- ions in the coagulation bath of alginic acid can simultaneously solidify the spider silk protein and alginic acid to form a silk thread. The color fastness of the spun fibers is high, the toughness is better than that of the original fibers, and the color is bright and stable. BRIEF DESCRIPTION OF DRAWINGS

[0030] In order to more clearly illustrate the technical solutions in the specific embodiments of the present application or the prior art, the drawings required to be used in the specific embodiments or the prior art description will be briefly introduced as follows. Obviously, the drawings described in the following description are some embodiments of the present application, and other drawings can also be obtained by those skilled in the art without creative effort on the basis of these drawings.

[0031] Figure 1 is the eluate of different fusion proteins in Example 2 of the present application; Figure 2 is the SDS-Page diagram of single fusion coral protein in Example 2 of the present application; in the diagram, the precipitate lane corresponds to the precipitate obtained by centrifugation after cell disruption, and the protein in the supernatant is used for subsequent protein purification, and the precipitate part is removed; Figure 3 is the SDS-Page diagram of double fusion coral protein in Example 2 of the present application; in the diagram, the precipitate lane corresponds to the precipitate obtained by centrifugation after cell disruption, and the protein in the supernatant is used for subsequent protein purification, and the precipitate part is removed; Figure 4 is the actual picture of recombinant marine coral protein seaweed fiber in Example 5 of the present application; Figure 5 is the sample tensile curve diagram of different single-end fusion coral proteins of spider silk in Example 5 of the present application; Figure 6 is the sample tensile curve diagram of different double-end fusion coral proteins in Example 5 of the present application. DETAILED DESCRIPTION

[0032] In order to better further understand the present application, the following examples are provided, but the following examples do not constitute a limitation on the content and protection scope of the present application, and any product which is the same or similar to the present application obtained by anyone under the inspiration of the present application or by combining the present application with other prior art features falls within the protection scope of the present application.

[0033] The specific experimental steps or conditions not mentioned in the examples are operated according to the conventional experimental steps or conditions in the art. The reagents or instruments not mentioned by the manufacturer are conventional reagent products or instruments which can be obtained by market purchase.

[0034] The sequences involved in the present application are as follows: Repeat region: 2rep amino acid sequence (SEQ ID NO. 19): GRGQGGYGQGSGGNAAAAAAAAAAAAAAAGQGGQGGYGRQSQGAGSAAAAAAAAAAAAAAGSGQGGYG GQGQGGYGQS. GGTCGTGGTCAAGGTGGTTATGGCCAAGGCAGCGGTGGTAATGCAGCAGCAGCAGCAGCGGCAGCAGCGGC AGCAGCGGCGGCGGCAGCGGGTCAAGGTGGTCAAGGTGGCTATGGTCGTCAAAGCCAAGGTGCAGGTTCAGCG GC

[0035] 2rep DNA sequence (SEQ ID NO. 20): GCAGCAGCAGCGGCGGCAGCGGCAGCGGCGGCAGCGGGCTCAGGCCAAGGTGGTTATGGCGGCCAAGGTCAAG GTG GTTATGGCCAGAGC. PQSPYGPGPQGPGPQGPGPQGPSGPGPQRPQGPGPQGPYGPGGVSVVSATVSGPGPQGPSGPGPQGPY GPGPQGPGPQGPGPQ LPGPQGPSGPGPQGPYGPGPQGPGPQGPGPQGPSGPGPQRPQGPGPQGPYGPGGVSVVSA TV. CCGCAGAGTCCATATGGCCCTGGCCCACAAGGCCCTGGGCCGCAGGGTCCAGGTCCGCAAGGCCCGTC GGCCCCTCAGCGCCCTCAAGGTCCTGGCCCGCAGGGCCCATATGGGCCAGGTGGTGTCAGCGTTGTCTCG GCAACTGTGAGCGGCCCTGGGCCGCAGGGTCCGAGCGGCCCAGGTCCGCAGGGTCCGTATGGTCCGGGCCCT CAGG

[0036] D3r amino acid sequence (SEQ ID NO. 21): GTCCGGGCCCGCAGGGCCCGGGTCCGCAACTGCCGGGGCCTCAGGGCCCGTCCGGTCCGGGTCCGCAGGGCCCGTA TGGTCCGGGCCCGCAAGGCCCGGGTCCGCAAGGTCCTGGCCCACAGGGCCCGAGTGGGCCAGGGCCGCAACGTCCG CAAGGCCCGGGCCCGCAGGGCCCTTACGGCCCTGGCGGAGTTAGTGTCGTGAGCGCAACCGTG. GAGEGAGGYRRYGGYGDGGAAAAAAAAAAAAAAAGGTGGYYYEAGGAGAAAAGAAAASAAGAAAGGAR ​

[0037] D3r DNA sequence (SEQ ID NO. 22): ​ ​ ​ ​ ​ ​

[0038] DmarAmSp amino acid sequence (SEQ ID NO. 23): ​ RAGGYGYGAGGAAGAAAAAGAAGGAGTRNTAYNSVGNS.

[0039] DmarAmSp DNA sequence (SEQ ID NO. 24): GGTGCAGGCGAAGGTGCCGGCGGATATCGCCGTTATGGTGGATACGGTGATGGAGGCGCCGCTGCCGC GGCGGCAGCGGCAGCAGCAGCCGCGGCGGCGGCTGGCGGTACTGGAGGATACTATTATGAAGCGGGTGGCGCCGGC GCAGCAGCGGCGGGTGCCGCTGCGGCATCCGCGGCCGGTGCAGCGGCAGGCGGTGCCCGCCGCGCCGGTGGTTACG GCTATGGGGCAGGAGGTGCCGCAGGCGCGGCGGCCGCAGCGGGTGCCGCCGGCGGTGCTGGCACCCGTAACACCGC GTATAATAGCGTGGGCAACTCG.

[0040] Flag amino acid sequence (SEQ ID NO. 25): VTVSSTVSVGGAGGPGAGGVGLGGAGAGGVGPGGFGGPGGFGGRGGPGGPGGPGGAGGGAGGAGGLYG PGGLYGPGGLYGPGGAGVPGAPGAPGAPGRPGGIGGGAGAGGVGPGGLSGGVGGSGVSVTESVTV.

[0041] Flag DNA sequence (SEQ ID NO. 26): GTAACTGTGTCGTCGACGGTTAGTGTCGGAGGAGCGGGCGGTCCCGGAGCAGGCGGTGTCGGTCTAGG TGGTGCGGGTGCAGGCGGCGTAGGTCCAGGTGGTTTCGGCGGACCGGGCGGCTTTGGCGGTCGTGGTGGCCCGGGC GGTCCAGGTGGTCCGGGTGGTGCCGGTGGTGGTGCCGGTGGCGCAGGTGGTCTGTATGGTCCAGGCGGTTTGTACG GGCCGGGGGGACTTTATGGCCCGGGCGGTGCTGGCGTGCCAGGTGCCCCGGGTGCCCCTGGTGCACCTGGCCGCCC TGGCGGCATTGGCGGCGGCGCCGGAGCGGGCGGCGTTGGCCCAGGTGGATTATCTGGTGGTGTCGGCGGCTCAGGT GTCTCGGTTACCGAAAGCGTTACCGTG.

[0042] EV1 amino acid sequence (SEQ ID NO. 27): GSAAAAAAAAAEAAAAAAAAAAAAGSGAGAGGAGGYGAGAGAGAGAGAGGAAGAGGAGGAGGAGGYGG ASVVYVGGGGAGAGAGSGAGAGSGAGAGAGSGAGAGGAGAAAGAGAGAGSGAGSGSGAGAGAGSGAGAGSGAGAGA GAGSGAAGGAGAGAGAGAGGS.

[0043] EV1 amino acid sequence (SEQ ID NO. 28): GGCTCCGCTGCTGCTGCGGCTGCTGCGGCTGCTGAAGCTGCTGCTGCTGCGGCAGCTGCTGCTGCTGC TGCTGGTTCTGGTGCAGGTGCCGGTGGTGCAGGTGGTTATGGTGCTGGTGCTGGTGCAGGCGCTGGTGCTGGTGCT GGTGGTGCAGCAGGTGCAGGTGGTGCAGGCGGTGCTGGTGGTGCTGGTGGTTACGGCGGCGCAAGCGTAGTATACGTTGGTGGCGGTGGTGCAGGTGCTGGTGCTGGTTCTGGTGCTGGTGCAGGTTCTGGTGCGGGTGCCGGTGCAGGTTC TGGTGCAGGTGCAGGTGGTGCTGGTGCAGCAGCTGGTGCGGGTGCAGGTGCAGGTTCTGGTGCCGGTTCTGGTTCC GGTGCTGGTGCAGGTGCTGGCTCCGGTGCTGGTGCTGGTAGCGGTGCAGGTGCAGGTGCTGGCGCAGGTTCTGGTG CAGCAGGTGGTGCTGGCGCAGGTGCAGGTGCTGGTGCAGGTGGATCC.

[0044] Globin NT amino acid sequence (SEQ ID NO. 29): MSHTTPWTNPGLAENFMNSFMQGLSSMPGFTASQLDDMSTIAQSMVQSIQSLAAQGRTSPNKLQALNM AFASSMAEIAASEEGGGSLSTKTSSIASAMSNAFLQTTGVVNQPFINEITQLVSMFAQAGMNDVSA.

[0045] Globin NT DNA sequence (SEQ ID NO. 30): ATGAGCCATACCACCCCGTGGACCAATCCTGGTCTGGCGGAAAACTTTATGAACAGCTTTATGCAGGG CCTGAGCAGCATGCCTGGTTTTACCGCGAGCCAGCTGGATGATATGAGCACCATTGCGCAGAGCATGGTGCAGAGC ATTCAGAGCTTAGCGGCGCAAGGTCGCACCAGCCCGAATAAATTACAGGCGCTGAACATGGCATTTGCGAGCAGCA TGGCGGAAATTGCGGCGTCAGAAGAAGGTGGTGGTAGCCTGAGCACCAAAACCAGCAGCATTGCGAGCGCGATGAG CAATGCGTTTCTGCAAACCACCGGCGTTGTTAACCAGCCGTTCATCAACGAAATTACCCAGCTGGTGAGCATGTTT GCACAGGCGGGCATGAATGATGTGAGCGCG.

[0046] Globin CT amino acid sequence (SEQ ID NO. 31): VTSGGYGYGTSAAAGAGVAAGSYAGAVNRLSSAEAASRVSSNIAAIASGGASALPSVISNIYSGVVAS GVSSNEALIQALLELLSALVHVLSSASIGNVSSVGVDSTLNVVQDSVGQYVG.

[0047] Globin CT amino acid sequence (SEQ ID NO. 32): GTTACCAGCGGCGGCTATGGTTATGGTACCAGCGCAGCAGCAGGTGCAGGCGTTGCAGCAGGTTCATA TGCAGGTGCGGTTAACCGTTTAAGCAGCGCGGAAGCAGCGTCACGTGTGAGCAGCAACATTGCGGCGATTGCAAGC GGTGGTGCGTCAGCATTACCGAGCGTGATTAGCAACATTTATAGCGGCGTGGTGGCGTCAGGTGTTAGCTCAAACG AAGCGCTGATTCAGGCGTTACTGGAATTACTGAGCGCGCTGGTGCATGTTTTAAGCAGCGCGAGCATTGGCAATGT GAGCAGCGTGGGTGTTGATAGCACCCTGAACGTGGTGCAGGATAGCGTGGGTCAGTATGTGGGC.

[0048] Example 1 Design of spider silk single-end fusion coral protein Single fusion coral protein 01: comprising NT globin, spider silk repeat region protein 2 rep, CT globin, coral fluorescent protein sfGFP connected in turn, named NT-2Rep-CT-sfGFP, its amino acid sequence is SEQ ID NO. 1: MGHHHHHHMSHTTPWTNPGLAENFMNSFMQGLSSMPGFTASQLDDMSTIAQSMVQSIQSLAAQGRTSPN KLQALNMAFASSMAEIAASEEGGGSLSTKTSSIASAMSNAFLQTTGVVNQPFINEITQLVSMFAQAGMNDVSAGNSG RGQGGYGQGSGGNAAAAAAAAAAAAAAAGQGGQGGYGRQSQGAGSAAAAAAAAAAAAAAGSGQGGYGGQGQGGYGQS GNS VTSGGYGYGTSAAAGAGVAAGSYAGAVNRLSSAEAASRVSSNIAAIASGGASALPSVISNIYSGVVASG VSSNEALIQALLELLSALVHVLSSASIGNVSSVGVDSTLNVVQDSVGQYVGGNS MRKGEELFTGVVPILVELDGDVNGHKFSVRGEGEGDATNGKLTLKFICTTGKLPVPWPTLVTTLTYGVQCFARYPDHMKQHDFFKSAMPEGYVQERTISFKDDGTYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNFNSHNVYITADKQKNGIKANFKIRHNVEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSVLSKDPNEKRDHMVLLEFVTAAGITHGMDELYK In the above amino acid sequence, the underlined His tag, the single underlined spider silk NT globulin, the wavy underlined inter-domain linker, the double underlined spider silk repeat protein 2rep, the dot-short line underlined CT globulin, and the non-underlined coral fluorescent protein sfGFP.

[0049] The DNA sequence of NT-2Rep-CT-sfGFP is shown in SEQ ID NO. 2: ATGGGCCATCATCATCATCATCATATGAGCCATACCACCCCGTGGACCAATCCTGGTCTGGCGGAAAAC TTTATGAACAGCTTTATGCAGGGCCTGAGCAGCATGCCTGGTTTTACCGCGAGCCAGCTGGATGATATGAGCACCAT TGCGCAGAGCATGGTGCAGAGCATTCAGAGCTTAGCGGCGCAAGGTCGCACCAGCCCGAATAAATTACAGGCGCTGA ACATGGCATTTGCGAGCAGCATGGCGGAAATTGCGGCGTCAGAAGAAGGTGGTGGTAGCCTGAGCACCAAAACCAGC AGCATTGCGAGCGCGATGAGCAATGCGTTTCTGCAAACCACCGGCGTTGTTAACCAGCCGTTCATCAACGAAATTAC CCAGCTGGTGAGCATGTTTGCACAGGCGGGCATGAATGATGTGAGCGCGGGCAATAGCGGTCGTGGTCAAGGTGGTT ATGGCCAAGGCAGCGGTGGTAATGCAGCAGCAGCAGCGGCAGCAGCGGCAGCAGCGGCGGCGGCAGCGGGTCAAGGT GGTCAAGGTGGCTATGGTCGTCAAAGCCAAGGTGCAGGTTCAGCGGCGGCAGCAGCAGCGGCGGCAGCGGCAGCGGC GGCAGCGGGCTCAGGCCAAGGTGGTTATGGCGGCCAAGGTCAAGGTGGTTATGGCCAGAGCGGTAACAGCGTTACCA GCGGCGGCTATGGTTATGGTACCAGCGCAGCAGCAGGTGCAGGCGTTGCAGCAGGTTCATATGCAGGTGCGGTTAAC CGTTTAAGCAGCGCGGAAGCAGCGTCACGTGTGAGCAGCAACATTGCGGCGATTGCAAGCGGTGGTGCGTCAGCATT ACCGAGCGTGATTAGCAACATTTATAGCGGCGTGGTGGCGTCAGGTGTTAGCTCAAACGAAGCGCTGATTCAGGCGT TACTGGAATTACTGAGCGCGCTGGTGCATGTTTTAAGCAGCGCGAGCATTGGCAATGTGAGCAGCGTGGGTGTTGAT AGCACCCTGAACGTGGTGCAGGATAGCGTGGGTCAGTATGTGGGCGGTAACAGCATGCGTAAAGGCGAAGAGCTGTTCACTGGTGTCGTCCCTATTCTGGTGGAACTGGATGGTGATGTCAACGGTCATAAGTTTTCCGTGCGTGGCGAGGGTGAAGGTGACGCAACTAATGGTAAACTGACGCTGAAGTTCATCTGTACTACTGGTAAACTGCCGGTACCTTGGCCGACTCTGGTAACGACGCTGACTTATGGTGTTCAGTGCTTTGCTCGTTATCCGGACCATATGAAGCAGCATGACTTCTTCAAGTCCGCCATGCCGGAAGGCTATGTGCAGGAACGCACGATTTCCTTTAAGGATGACGGCACGTACAAAACGCGTGCGGAAGTGAAATTTGAAGGCGATACCCTGGTAAACCGCATTGAGCTGAAAGGCATTGACTTTAAAGAAGACGGCAATATCCTGGGCCATAAGCTGGAATACAATTTTAACAGCCACAATGTTTACATCACCGCCGATAAACAAAAAAATGGCATTAAAGCGAATTTTAAAATTCGCCACAACGTGGAGGATGGCAGCGTGCAGCTGGCTGATCACTACCAGCAAAACACTCCAATCGGTGATGGTCCTGTTCTGCTGCCAGACAATCACTATCTGAGCACGCAAAGCGTTCTGTCTAAAGATCCGAACGAGAAACGCGATCATATGGTTCTGCTGGAGTTCGTAACCGCAGCGGGCATCACGCATGGTATGGATGAACTGTACAAATAA In the above nucleotide sequence, the dotted underlined is His tag, the single underlined is NT globulin of spider silk, the wavy underlined is Linker between regions, the double underlined is spider silk repeat region protein 2rep, the dot-short line underlined is CT globulin, and the non-underlined is coral fluorescent protein sfGFP.

[0050] Single fusion coral protein 02: comprising coral fluorescent protein sfGFP, NT globulin, spider silk repeat region protein 2rep, and CT globulin connected in turn, named sfGFP-NT-2Rep-CT, and its amino acid sequence is shown in SEQ ID NO. 3: MGHHHHHHMRKGEELFTGVVPILVELDGDVNGHKFSVRGEGEGDATNGKLTLKFICTTGKLPVPWPTLVTTLTYGVQCFARYPDHMKQHDFFKSAMPEGYVQERTISFKDDGTYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNFNSHNVYITADKQKNGIKANFKIRHNVEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSVLSKDPNEKRDHMVLLEFVTAAGITHGMDELYK GNSMSHTTPWTNPGLAENFMNSFMQGLSSMPGFTASQLDDMSTIAQSMVQ SIQSLAAQGRTSPNKLQALNMAFASSMAEIAASEEGGGSLSTKTSSIASAMSNAFLQTTGVVNQPFINEITQLVSM FAQAGMNDVSAGNSGRGQGGYGQGSGGNAAAAAAAAAAAAAAAGQGGQGGYGRQSQGAGSAAAAAAAAAAAAAAGS GQGGYGGQGQGGYGQSGNSVTSGGYGYGTSAAAGAGVAAGSYAGAVNRLSSAEAASRVSSNIAAIASGGASALPSV ISNIYSGVVASGVSSNEALIQALLELLSALVHVLSSASIGNVSSVGVDSTLNVVQDSVGQYVG* In the above sequence, the underlined dash line is His tag, the non-underlined is sfGFP, the underlined wave line is Linker between regions, the single underlined is NT globulin of spider silk, the double underlined is 2Rep of spider silk repeat region, and the underlined dot-short line is CT globulin.

[0051] The DNA sequence of sfGFP-NT-2Rep-CT is shown in SEQ ID NO. 4: ATGGGCCATCATCATCATCATCATATGCGTAAAGGCGAAGAGCTGTTCACTGGTGTCGTCCCTATTCTGGTGGAACTGGATGGTGATGTCAACGGTCATAAGTTTTCCGTGCGTGGCGAGGGTGAAGGTGACGCAACTAATGGTAAACTGACGCTGAAGTTCATCTGTACTACTGGTAAACTGCCGGTACCTTGGCCGACTCTGGTAACGACGCTGACTTATGGTGTTCAGTGCTTTGCTCGTTATCCGGACCATATGAAGCAGCATGACTTCTTCAAGTCCGCCATGCCGGAAGGCTATGTGCAGGAACGCACGATTTCCTTTAAGGATGACGGCACGTACAAAACGCGTGCGGAAGTGAAATTTGAAGGCGATACCCTGGTAAACCGCATTGAGCTGAAAGGCATTGACTTTAAAGAAGACGGCAATATCCTGGGCCATAAGCTGGAATACAATTTTAACAGCCACAATGTTTACATCACCGCCGATAAACAAAAAAATGGCATTAAAGCGAATTTTAAAATTCGCCACAACGTGGAGGATGGCAGCGTGCAGCTGGCTGATCACTACCAGCAAAACACTCCAATCGGTGATGGTCCTGTTCTGCTGCCAGACAATCACTATCTGAGCACGCAAAGCGTTCTGTCTAAAGATCCGAACGAGAAACGCGATCATATGGTTCTGCTGGAGTTCGTAACCGCAGCGGGCATCACGCATGGTATGGATGAACTGTACAAA GGTAACAGCATGAGCCATACCACC CCGTGGACCAATCCTGGTCTGGCGGAAAACTTTATGAACAGCTTTATGCAGGGCCTGAGCAGCATGCCTGGTTTTAC CGCGAGCCAGCTGGATGATATGAGCACCATTGCGCAGAGCATGGTGCAGAGCATTCAGAGCTTAGCGGCGCAAGGTC GCACCAGCCCGAATAAATTACAGGCGCTGAACATGGCATTTGCGAGCAGCATGGCGGAAATTGCGGCGTCAGAAGAA GGTGGTGGTAGCCTGAGCACCAAAACCAGCAGCATTGCGAGCGCGATGAGCAATGCGTTTCTGCAAACCACCGGCGT TGTTAACCAGCCGTTCATCAACGAAATTACCCAGCTGGTGAGCATGTTTGCACAGGCGGGCATGAATGATGTGAGCG CGGGCAATAGCGGTCGTGGTCAAGGTGGTTATGGCCAAGGCAGCGGTGGTAATGCAGCAGCAGCAGCGGCAGCAGCG GCAGCAGCGGCGGCGGCAGCGGGTCAAGGTGGTCAAGGTGGCTATGGTCGTCAAAGCCAAGGTGCAGGTTCAGCGGC GGCAGCAGCAGCGGCGGCAGCGGCAGCGGCGGCAGCGGGCTCAGGCCAAGGTGGTTATGGCGGCCAAGGTCAAGGTG GTTATGGCCAGAGCGGTAACAGCGTTACCAGCGGCGGCTATGGTTATGGTACCAGCGCAGCAGCAGGTGCAGGCGTT GCAGCAGGTTCATATGCAGGTGCGGTTAACCGTTTAAGCAGCGCGGAAGCAGCGTCACGTGTGAGCAGCAACATTGC GGCGATTGCAAGCGGTGGTGCGTCAGCATTACCGAGCGTGATTAGCAACATTTATAGCGGCGTGGTGGCGTCAGGTG TTAGCTCAAACGAAGCGCTGATTCAGGCGTTACTGGAATTACTGAGCGCGCTGGTGCATGTTTTAAGCAGCGCGAGC ATTGGCAATGTGAGCAGCGTGGGTGTTGATAGCACCCTGAACGTGGTGCAGGATAGCGTGGGTCAGTATGTGGGC In the above sequence, the dotted underlined is His tag, the non-underlined is coral fluorescent protein sfGFP, the wavy underlined is the Linker between regions, the single underlined is NT globulin of spider silk, the double underlined is spider silk repeat region protein 2rep, and the dot-short line underlined is CT globulin.

[0052] Single fusion coral protein 03: comprising NT globulin, spider silk repeat region protein 2rep, CT globulin, and coral fluorescent protein eforRed connected in turn, named NT-2Rep-eforRed, and its amino acid sequence is shown in SEQ ID NO. 5: MGHHHHHHMSHTTPWTNPGLAENFMNSFMQGLSSMPGFTASQLDDMSTIAQSMVQSIQSLAAQGRTSP NKLQALNMAFASSMAEIAASEEGGGSLSTKTSSIASAMSNAFLQTTGVVNQPFINEITQLVSMFAQAGMNDVSAGN SGRGQGGYGQGSGGNAAAAAAAAAAAAAAAGQGGQGGYGRQSQGAGSAAAAAAAAAAAAAAGSGQGGYGGQGQGGY GQSGNS MSVIKQVMKTKLHLEGTVNGHDFTIEGKGEGKPYEGLQHMKMTVTKGAPLPFSVHILTPSHMYGSKPFNKYPADIPDYHKQSFPEGMSWERSMIFEDGGVCTASNHSSINLQENCFIYDVKFHGVNLPPDGPVMQKTIAGWEPSVETLYVRDGMLKSDTAMVFKLKGGGHHRVDFKTTYKAKKPVKLPEFHFVEHRLELTKHDKDFTTWDQQEAAEGHFSPLPKALP In the above sequence, the dotted underlined is His tag, the single underlined is NT globulin of spider silk, the wavy underlined is the Linker between regions, the double underlined is spider silk repeat region protein 2rep, and the non-underlined is coral fluorescent protein eforRed.

[0053] The DNA sequence of NT-2Rep-eforRed is shown in SEQ ID NO. 6: ATGGGCCATCATCATCATCATCATATGAGCCATACCACCCCGTGGACCAATCCTGGTCTGGCGGAAAAC TTTATGAACAGCTTTATGCAGGGCCTGAGCAGCATGCCTGGTTTTACCGCGAGCCAGCTGGATGATATGAGCACCAT TGCGCAGAGCATGGTGCAGAGCATTCAGAGCTTAGCGGCGCAAGGTCGCACCAGCCCGAATAAATTACAGGCGCTGA ACATGGCATTTGCGAGCAGCATGGCGGAAATTGCGGCGTCAGAAGAAGGTGGTGGTAGCCTGAGCACCAAAACCAGC AGCATTGCGAGCGCGATGAGCAATGCGTTTCTGCAAACCACCGGCGTTGTTAACCAGCCGTTCATCAACGAAATTAC CCAGCTGGTGAGCATGTTTGCACAGGCGGGCATGAATGATGTGAGCGCGGGCAATAGCGGTCGTGGTCAAGGTGGTT ATGGCCAAGGCAGCGGTGGTAATGCAGCAGCAGCAGCGGCAGCAGCGGCAGCAGCGGCGGCGGCAGCGGGTCAAGGT GGTCAAGGTGGCTATGGTCGTCAAAGCCAAGGTGCAGGTTCAGCGGCGGCAGCAGCAGCGGCGGCAGCGGCAGCGGC GGCAGCGGGCTCAGGCCAAGGTGGTTATGGCGGCCAAGGTCAAGGTGGTTATGGCCAGAGCGGTAACAGCATGTCAGTGATTAAGCAGGTAATGAAGACCAAGTTGCACCTTGAGGGCACTGTCAATGGCCATGATTTTACGATCGAGGGTAAAGGTGAAGGCAAGCCGTACGAAGGGTTACAGCACATGAAAATGACAGTCACCAAAGGCGCGCCTCTGCCGTTTTCCGTTCATATTCTTACACCTAGCCACATGTATGGAAGCAAACCGTTTAATAAGTATCCAGCGGATATCCCAGACTACCACAAACAGTCTTTTCCCGAAGGTATGTCTTGGGAGCGGTCGATGATTTTTGAAGATGGTGGCGTATGCACCGCCAGTAATCACTCCAGCATAAACTTGCAAGAGAACTGTTTCATCTATGATGTTAAATTTCATGGTGTGAACCTGCCTCCGGATGGGCCCGTAATGCAAAAAACCATTGCTGGATGGGAGCCGAGCGTGGAAACACTGTACGTGCGTGACGGGATGTTAAAAAGTGACACTGCAATGGTTTTTAAACTGAAAGGAGGCGGTCATCATCGTGTTGATTTCAAAACGACGTATAAAGCCAAAAAACCTGTCAAGCTGCCAGAATTTCATTTCGTTGAACATCGCCTGGAACTGACCAAACACGATAAAGATTTCACAACTTGGGACCAGCAGGAGGCAGCCGAAGGCCATTTCTCACCGCTGCCGAAGGCTCTCCCA The above sequence has a His tag underlined with a dotted line, a spider silk NT globulin underlined with a single line, a linker underlined with a wavy line between regions, a spider silk repeat protein 2 rep underlined with a double line, and a coral fluorescent protein eforRed without underlining. The CT globulin is underlined with a dot-short line.

[0054] Single fusion coral protein 04: comprising a coral fluorescent protein amilCP, a spider silk repeat protein 2 rep, and a CT globulin connected in sequence, named amilCP-2Rep-CT, and its amino acid sequence is shown in SEQ ID NO. 7: MGHHHHHHMSVIAKQMTYKVYMSGTVNGHYFEVEGDGKGKPYEGEQTVKLTVTKGGPLPFAWDILSPQCQYGSIPFTKYPEDIPDYVKQSFPEGYTWERIMNFEDGAVCTVSNDSSIQGNCFIYHVKFSGLNFPPNGPVMQKKTQGWEPNTERLFARDGMLLGNNFMALKLEGGGHYLCEFKTTYKAKKPVKMPGYHYVDRKLDVTNHNKDYTSVEQCEISIARKPVVA GNSGRGQGGYGQGSGGNAAAAAAAAAAAAAAAGQGGQGGYGRQSQGAGSAAAAAAAAAAAAAAGSGQ GGYGGQGQGGYGQSGNSVTSGGYGYGTSAAAGAGVAAGSYAGAVNRLSSAEAASRVSSNIAAIASGGASALPSVIS NIYSGVVASGVSSNEALIQALLELLSALVHVLSSASIGNVSSVGVDSTLNVVQDSVGQYVG* In the above sequence, the underlined dash line is His tag, the non-underlined is amilCP of coral fluorescent protein, the underlined wave is Linker between regions, the double underlined is Rep of Repilin-2, and the underlined dash-dot is CT globulin.

[0055] The DNA sequence of amilCP-2Rep-CT is shown in SEQ ID NO. 8: ATGGGCCATCATCATCATCATCATATGAGTGTGATCGCTAAACAAATGACCTACAAGGTTTATATGTCAGGCACGGTCAATGGACACTACTTTGAGGTCGAAGGCGATGGAAAAGGTAAGCCCTACGAGGGGGAGCAGACGGTAAAGCTCACTGTCACCAAGGGCGGACCTCTGCCATTTGCTTGGGATATTTTATCACCACAGTGTCAGTACGGAAGCATACCATTCACCAAGTACCCTGAAGACATCCCTGACTATGTAAAGCAGTCATTCCCGGAGGGCTATACATGGGAGAGGATCATGAACTTTGAAGATGGTGCAGTGTGTACTGTCAGCAATGATTCCAGCATCCAAGGCAACTGTTTCATCTACCATGTCAAGTTCTCTGGTTTGAACTTTCCTCCCAATGGACCTGTCATGCAGAAGAAGACACAGGGCTGGGAACCCAACACTGAGCGTCTCTTTGCACGAGATGGAATGCTGCTAGGAAACAACTTTATGGCTCTGAAGTTAGAAGGAGGCGGTCACTATTTGTGTGAATTTAAAACTACTTACAAGGCAAAGAAGCCTGTGAAGATGCCAGGGTATCACTATGTTGACCGCAAACTGGATGTAACCAATCACAACAAGGATTACACTTCGGTTGAGCAGTGTGAAATTTCCATTGCACGCAAACCTGTGGTCGCC GGCAATAGCGGTCGTGGTCAAGGTGGTTATGGCCAAGGCAGCGGTGGTAATGCAGCAGCAGCAGCGGCAGCAGCG GCAGCAGCGGCGGCGGCAGCGGGTCAAGGTGGTCAAGGTGGCTATGGTCGTCAAAGCCAAGGTGCAGGTTCAGCGGC GGCAGCAGCAGCGGCGGCAGCGGCAGCGGCGGCAGCGGGCTCAGGCCAAGGTGGTTATGGCGGCCAAGGTCAAGGTG GTTATGGCCAGAGCGGTAACAGCGTTACCAGCGGCGGCTATGGTTATGGTACCAGCGCAGCAGCAGGTGCAGGCGTT GGCGATTGCAAGCGGTGGTGCGTCAGCATTACCGAGCGTGATTAGCAACATTTATAGCGGCGTGGTGGCGTCAGGTG TTAGCTCAAACGAAGCGCTGATTCAGGCGTTACTGGAATTACTGAGCGCGCTGGTGCATGTTTTAAGCAGCGCGAGC ATTGGCAATGTGAGCAGCGTGGGTGTTGATAGCACCCTGAACGTGGTGCAGGATAGCGTGGGTCAGTATGTGGGCTA Figure 1 A In the above sequence, the dotted underlined is His tag, the non-underlined is coral fluorescent protein amilCP, the wavy underlined is Linker between regions, the double underlined is spider silk repeat region protein 2rep, and the dot-short line underlined is CT globulin.

[0056] The above sequences are synthesized by silicon-based biosynthesis, and the above synthetic sequences are inserted into a pET28a commercial plasmid vector, the insertion site is between BamHI and XhoI, and finally the plasmid is transformed into Escherichia coli BL21 (DE3) strain by chemical transformation method (completed by silicon-based biosynthesis service).

[0057] Example 2 Expression and purification of spider silk single-end fusion coral protein The constructed strain was fermented, purified and freeze-dried to obtain a recombinant protein dry powder, as follows: (1) Shake flask fermentation of bacterial cells The constructed strain of Example 1 was inoculated on a solid LB (containing 50 ug / ml kanamycin Kana) plate with preserved glycerol bacteria, and incubated in a 37°C incubator overnight. A single colony was picked and inoculated into 4-5 mL of LB liquid medium (containing 50 ug / ml kanamycin Kana), and incubated at 37°C, 200 rpm overnight. The next day, 1% of the volume was inoculated into a 400 mL shake flask, and incubated at 200 rpm, 37°C for 2-3 h until the OD 600 =0.6~0.8. Isopropyl thiogalactoside (IPTG) was added to a final concentration of 0.3 mM, and the culture was induced at 20°C for 12-16 h. The bacterial cells were collected by centrifugation at 12000 rpm for 5 min.

[0058] (2) Protein purification The bacterial cells were resuspended in 20 mM Tris-HCl to a concentration of OD 600 =20 or so. Ultrasonic treatment was performed at 25 kHz, power 70-80%, 3 s on and 6 s off, for a total of 30 min, until the solution changed from turbid to clear. The supernatant was collected by centrifugation at 12000 rpm for 10-15 min. The supernatant was passed through a pre-packed nickel column (Shanghai Generay Biotech Co., Ltd., product number NO. A600657) 2-3 times, and the flow-through was collected. The column was washed with 20 mM Tris-HCl 3-5 times (1-3 times the column volume), until the eluted solution was clear. The column was washed with 1-2 times the column volume of 0.5 M imidazole solution at least 3 times, and the eluate was collected. The remaining 0.5 M imidazole solution was removed by washing the column with 5 times the column volume of 20 mM Tris-HCl, and the column was finally stored in 20% alcohol. The purified protein was dialyzed in water. Figure 2 The color of different protein solutions was different, as shown in the figure.

[0059] (3) SDS-PAGE electrophoresis A 10% SDS-PAGE gel (Yaenzyme, product number specification: PG212) was prepared.

[0060] Sample preparation and detection: 20 uL of the sample from the protein purification process was added to 5 uL of loading buffer, heated at 98°C for 15 min to denature the protein, and then 5 uL was taken for SDS-PAGE detection. The electrophoresis detection results are shown in the figure. Figure 3 andMSHSKQALADTMKMTWLMEGSVNGHAFTIEGEGTGKPYEGKQSGTFRVTKGGPLPFAFDIVAPTLKYG as shown.

[0061] (4) Protein freeze-drying After dialysis, the protein solution was placed in a -80°C refrigerator until the protein solution completely solidified, and then placed in a freeze dryer for freeze-drying, with a freezing temperature of -55°C and a vacuum degree of 5 Pa. The freeze-dried protein was weighed, and finally the purified recombinant coral protein was obtained.

[0062] Example 3 Design of recombinant double-end fusion coral protein Double fusion coral protein 01: containing coral color protein spisPink, spider silk repeat region protein 2 rep, and coral fluorescent protein mScarlet3 connected in turn, named spisPink-2Rep-mScarlet3, the amino acid sequence is SEQ ID NO. 9: FKCFMKYPADIPDYFKLAFPEGLTYDRKIAFEDGGCATATVEMSLKGNTLVHKTNFQGGNFPIDGPVMQKRTLGWE PTSEKMTPCDGIIKGDTIMYLMVEGGKTLKCRYENNYRANKPVLMPPSHFVDLRLTRTNLDKEGLAFKLEEYAVAR VLEVGSSSGSSSGNSGRGQGGYGQGSGGNAAAAAAAAAAAAAAAGQGGQGGYGRQSQGAGSAAAAAAAAAAAAAAG SGQGGYGGQGQGGYGQSGNS HHHHHH* MDSTEAVIKEFMRFKVHMEGSMNGHEFEIEGEGEGRPYEGTQTAKLKVTKGGPLPFSWDILSPQFMYGSRAFIKHPADIPDYWKQSFPEGFKWERVMIFEDGGTVSVTQDTSLEDGTLIYKVKLRGGNFPPDGPVMQKRTMGWEASTERLYPEDVVLKGDIKMALRLKDGGRYLADFKTTYKAKKPVQMPGAFNIDRKLDITSHNEDYTVVEQYERSVARHSTGGSGGS ATGTCGCACTCAAAACAAGCACTGGCAGATACGATGAAAATGACGTGGCTGATGGAAGGTTCGGTCAAC In the above sequence, the single underline is the coral color protein spisPink, the wavy underline is the Linker between regions, the double underline is the spider silk repeat region protein 2 rep, the non-underlined is the coral fluorescent protein mScarlet3, and the dotted underline is the His tag.

[0063] The DNA sequence of spisPink-2Rep-mScarlet3 is SEQ ID NO. 10: GGTCACGCCTTTACGATTGAAGGCGAAGGTACCGGCAAACCGTATGAAGGTAAACAGAGTGGCACCTTTCGTGTTAC GAAAGGCGGTCCGCTGCCGTTTGCGTTTGATATTGTCGCTCCGACCCTGAAATACGGTTTCAAATGCTTCATGAAAT ACCCGGCGGATATTCCGGACTACTTTAAACTGGCCTTCCCGGAAGGCCTGACGTACGATCGTAAAATTGCGTTTGAA ​ GACGGCGGTTGTGCGACCGCCACGGTCGAAATGAGCCTGAAGGGTAACACCCTGGTGCATAAAACGAACTTTCAGGG CGGTAATTTCCCGATTGATGGTCCGGTGATGCAAAAACGCACCCTGGGCTGGGAACCGACCTCTGAAAAAATGACGC CGTGCGATGGTATTATCAAAGGCGACACCATCATGTACCTGATGGTTGAAGGCGGTAAAACGCTGAAATGTCGTTAT GAAAACAATTACCGCGCCAACAAACCAGTGCTGATGCCGCCGAGCCACTTTGTGGATCTGCGTCTGACCCGCACGAA TCTGGATAAAGAAGGTCTGGCGTTCAAACTGGAAGAATATGCTGTTGCCCGTGTGCTGGAAGTGGGCTCAAGCTCCG GAAGTAGCAGTGGCAATAGCGGTCGTGGTCAAGGTGGTTATGGCCAAGGCAGCGGTGGTAATGCAGCAGCAGCAGCG GCAGCAGCGGCAGCAGCGGCGGCGGCAGCGGGTCAAGGTGGTCAAGGTGGCTATGGTCGTCAAAGCCAAGGTGCAGG TTCAGCGGCGGCAGCAGCAGCGGCGGCAGCGGCAGCGGCGGCAGCGGGCTCAGGCCAAGGTGGTTATGGCGGCCAAG GTCAAGGTGGTTATGGCCAGAGCGGTAACAGCATGGACTCCACCGAGGCTGTTATCAAAGAATTCATGCGTTTCAAAGTGCACATGGAAGGTTCTATGAACGGTCACGAATTCGAGATCGAAGGTGAAGGCGAAGGTCGTCCGTACGAAGGCACCCAGACCGCCAAACTGAAAGTTACCAAAGGTGGCCCGCTGCCATTCAGCTGGGATATCCTGAGCCCGCAGTTCATGTATGGTAGCCGCGCGTTTATCAAACACCCGGCGGACATCCCAGACTACTGGAAACAATCCTTCCCTGAAGGCTTTAAATGGGAACGTGTGATGATCTTCGAGGACGGCGGTACCGTTAGCGTGACGCAAGATACTAGCCTGGAGGACGGTACCCTGATCTACAAAGTTAAGCTGCGTGGCGGTAATTTTCCGCCAGACGGTCCGGTAATGCAGAAACGTACCATGGGTTGGGAAGCGTCTACCGAACGTCTGTACCCAGAAGATGTAGTTCTGAAAGGCGACATTAAAATGGCGCTGCGTCTGAAGGATGGTGGTCGTTACCTGGCAGACTTCAAAACTACGTACAAGGCAAAAAAACCGGTCCAGATGCCGGGCGCATTCAACATCGACCGCAAACTGGACATCACCTCTCACAACGAAGACTATACCGTAGTTGAGCAGTACGAACGTTCCGTAGCACGCCACAGCACTGGTGGCTCTGGTGGCTCC CATCATCATCACCATCATTAA In the above sequence, the single underlined sequence is the coral protein spisPink, the wavy underlined sequence is the linker between regions, the double underlined sequence is the spider silk repeat region protein 2rep, the non-underlined sequence is the coral fluorescent protein mScarlet3, and the dotted underlined sequence is the His tag.

[0064] The double fusion coral protein 02 comprises the coral protein gfasPurple, the spider silk repeat region protein D3r and the coral fluorescent protein mScarlet3 connected in sequence, and is named gfasPurple-D3r-mScarlet3. The amino acid sequence is shown in SEQ ID NO. 11: MSVIAKQMTYKVYMSGTVNGHYFEVEGDGKGKPYEGEQTVKLTVTKGGPLPFAWDILSPQSQYGSIPF TKYPEDIPDYVKQSFPEGYTWERIMNFEDGAVCTVSNDSSIQGNCFIYHVKFSGLNFPPNGPVMQKKTQGWEPNTE RLFARDGMLIGNNFMALKLEGGGHYLCEFKSTYKAKKPVKMPGYHYVDRKLDVTNHNKDYTSVEQCEISIARKSVV AGSSSGSSSGSPQSPYGPGPQGPGPQGPGPQGPSGPGPQRPQGPGPQGPYGPGGVSVVSATVSGPGPQGPSGPGPQGPYGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGPQGPGP ​ MDSTEAVIKEFMRFKVHMEGSMNGHEFEIEGEGEGRPYEGTQTAKLKVTKGGPLPFSWDILSPQFMYGSRAFIKHPADIPDYWKQSFPEGFKWERVMIFEDGGTVSVTQDTSLEDGTLIYKVKLRGGNFPPDGPVMQKRTMGWEASTERLYPEDVVLKGDIKMALRLKDGGRYLADFKTTYKAKKPVQMPGAFNIDRKLDITSHNEDYTVVEQYERSVARHSTGGSGGS ​ In the above sequence, the single underline is the coral fluorescent protein gfasPurple, the wavy underline is the inter-region linker, the double underline is the spider silk repeat region protein D3r, the no underline is the coral fluorescent protein mScarlet3, and the dotted underline is the His tag.

[0065] The DNA sequence of gfasPurple-D3r-mScarlet3 is shown in SEQ ID NO. 12: ​ ​ ​ ​ ​ ​ ​ ​ ​ ​ ​ AACTGTGAGCGGCCCTGGGCCGCAGGGTCCGAGCGGCCCAGGTCCGCAGGGTCCGTATGGTCCGGGCCCTCAGGGTC CGGGCCCGCAGGGCCCGGGTCCGCAACTGCCGGGGCCTCAGGGCCCGTCCGGTCCGGGTCCGCAGGGCCCGTATGGT CCGGGCCCGCAAGGCCCGGGTCCGCAAGGTCCTGGCCCACAGGGCCCGAGTGGGCCAGGGCCGCAACGTCCGCAAGG CCCGGGCCCGCAGGGCCCTTACGGCCCTGGCGGAGTTAGTGTCGTGAGCGCAACCGTGTCTGGATCCATGGACTCCACCGAGGCTGTTATCAAAGAATTCATGCGTTTCAAAGTGCACATGGAAGGTTCTATGAACGGTCACGAATTCGAGATCGAAGGTGAAGGCGAAGGTCGTCCGTACGAAGGCACCCAGACCGCCAAACTGAAAGTTACCAAAGGTGGCCCGCTGCCATTCAGCTGGGATATCCTGAGCCCGCAGTTCATGTATGGTAGCCGCGCGTTTATCAAACACCCGGCGGACATCCCAGACTACTGGAAACAATCCTTCCCTGAAGGCTTTAAATGGGAACGTGTGATGATCTTCGAGGACGGCGGTACCGTTAGCGTGACGCAAGATACTAGCCTGGAGGACGGTACCCTGATCTACAAAGTTAAGCTGCGTGGCGGTAATTTTCCGCCAGACGGTCCGGTAATGCAGAAACGTACCATGGGTTGGGAAGCGTCTACCGAACGTCTGTACCCAGAAGATGTAGTTCTGAAAGGCGACATTAAAATGGCGCTGCGTCTGAAGGATGGTGGTCGTTACCTGGCAGACTTCAAAACTACGTACAAGGCAAAAAAACCGGTCCAGATGCCGGGCGCATTCAACATCGACCGCAAACTGGACATCACCTCTCACAACGAAGACTATACCGTAGTTGAGCAGTACGAACGTTCCGTAGCACGCCACAGCACTGGTGGCTCTGGTGGCTCC CATCATCATCACCATC ATTAA In the above sequence, the single underlined sequence is the coral fluorescent protein gfasPurple, the wavy underlined sequence is the inter-region linker, the double underlined sequence is the spider silk repeat region protein D3r, the non-underlined sequence is the coral fluorescent protein mScarlet3, and the dotted underlined sequence is the His tag.

[0066] Double fusion coral protein 03: comprising the coral fluorescent protein amilCP orange, the spider silk repeat region protein DmarAmSp, and the coral fluorescent protein FwYellow connected in sequence, named amilCP orange-Dmar-FwYellow, and its amino acid sequence is shown in SEQ ID NO. 13: MSVIAKQMTYKVYMSGTVNGHYFEVEGDGKGKPYEGEQTVKLTVTKGGPLPFAWDILSPQVGYGSIPF TKYPEDIPDYVKQSFPEGYTWERIMNFEDGAVCTVSNDSSIQGNCFIYHVKFSGLNFPPNGPVMQKKTQGWEPNTERLFARDGMLLGNNFMALKLEGGGHYLCEFKTTYKAKKPVKMPGYHYVDRKLDVTNHNKDYTSVEQCEISIARKPVV AGSSSGSSSGNSGAGEGAGGYRRYGGYGDGGAAAAAAAAAAAAAAAGGTGGYYYEAGGAGAAAAGAAAASAAGAAA GGARRAGGYGYGAGGAAGAAAAAGAAGGAGTRNTAYNSVGNSSGS MTALTEGAKLFEKEIPYITELEGDVEGMKFIIKGEGTGDASVGKVDAQFICTTGDVPVPWSTLVTTLTYGAQCFAKYPRHIADFFKSCMPEGYVQERTITFEGDGVFKTRAEVTFENGSVYNRVKLNGQGFKKDGHVLGKNLEFNFTPHCLYIWGDQANHGLKSAFKIMHEITGSKEDFIVADHTQMNTPIGGGPVHVPEYHHITYHVTLSKDVTDHRDHLNIVEVIKAVDLETYR HHHHHH* Single underlined is amilCP orange, wavy underlined is the interdomain linker, double underlined is DmarAmSp, and un- underlined is FwYellow.

[0067] The DNA sequence of amilCP orange-Dmar-FwYellow is SEQ ID NO. 14: ATGTCTGTAATCGCAAAACAAATGACCTACAAAGTTTACATGTCTGGTACGGTTAACGGTCATTACTTC GAAGTCGAAGGCGATGGCAAAGGCAAACCGTATGAAGGTGAACAGACCGTGAAACTGACTGTAACCAAAGGTGGTCC TCTGCCATTTGCGTGGGATATTCTGTCTCCGCAGGTTGGATACGGTTCCATCCCGTTTACTAAATACCCGGAAGACA TCCCGGACTACGTAAAGCAGTCTTTTCCGGAGGGCTATACCTGGGAACGCATTATGAACTTCGAAGATGGCGCTGTG TGCACTGTGTCCAATGATTCCTCCATCCAGGGCAACTGTTTTATCTACCACGTTAAATTCAGCGGTCTGAATTTCCC GCCGAACGGTCCGGTCATGCAGAAAAAAACTCAGGGCTGGGAACCGAACACTGAACGTCTGTTCGCTCGTGATGGCA TGCTGCTGGGCAATAACTTCATGGCCCTGAAGCTGGAGGGTGGTGGTCACTACCTGTGTGAGTTCAAAACTACCTAC AAAGCGAAAAAGCCAGTGAAAATGCCAGGTTATCACTATGTGGATCGTAAACTGGATGTGACGAACCACAACAAAGA CTACACTAGCGTTGAACAGTGCGAAATCAGCATCGCTCGTAAGCCGGTAGTGGCGGGCTCAAGCTCCGGAAGTAGCA GTGGTAATAGTGGTGCAGGCGAAGGTGCCGGCGGATATCGCCGTTATGGTGGATACGGTGATGGAGGCGCCGCTGCC GCGGCGGCAGCGGCAGCAGCAGCCGCGGCGGCGGCTGGCGGTACTGGAGGATACTATTATGAAGCGGGTGGCGCCGG CGCAGCAGCGGCGGGTGCCGCTGCGGCATCCGCGGCCGGTGCAGCGGCAGGCGGTGCCCGCCGCGCCGGTGGTTACG GCTATGGGGCAGGAGGTGCCGCAGGCGCGGCGGCCGCAGCGGGTGCCGCCGGCGGTGCTGGCACCCGTAACACCGCG TATAATAGCGTGGGCAACTCGTCTGGATCCATGACGGCACTGACTGAAGGCGCAAAACTGTTCGAGAAAGAAATCCCATATATCACTGAGCTGGAAGGTGACGTTGAAGGTATGAAGTTTATCATCAAGGGTGAAGGTACCGGTGACGCGAGCGTCGGTAAAGTGGATGCTCAGTTCATTTGTACCACGGGCGACGTTCCGGTTCCGTGGAGCACGCTGGTCACCACGCTGACGTATGGTGCTCAGTGCTTTGCCAAGTATCCGCGCCACATTGCGGATTTCTTCAAAAGCTGCATGCCGGAAGGTTACGTCCAAGAGCGCACCATCACCTTTGAGGGTGATGGCGTGTTCAAGACCCGTGCGGAAGTCACCTTTGAAAATGGCAGCGTGTACAACCGTGTAAAACTGAACGGCCAGGGTTTCAAGAAGGACGGCCACGTGCTGGGCAAAAATCTGGAGTTTAACTTTACCCCTCATTGTTTGTACATTTGGGGTGACCAAGCGAATCATGGCCTGAAGAGCGCGTTCAAAATCATGCATGAGATCACCGGCTCCAAAGAGGATTTCATTGTTGCCGATCACACCCAAATGAATACCCCGATTGGTGGTGGTCCGGTGCACGTGCCGGAGTACCACCACATTACGTATCATGTTACCCTGTCTAAAGACGTCACCGATCACCGTGACCATTTGAACATTGTTGAGGTGATCAAGGCAGTTGACCTGGAGACGTACCGT CATCATCATCACCATCATTAA In the above sequence, the single underlined sequence is the coral protein amilCP orange, the wavy underlined sequence is the linker between regions, the double underlined sequence is the spider silk repeat region protein DmarAmSp, the non-underlined sequence is the coral fluorescent protein FwYellow, and the dotted underlined sequence is the His tag.

[0068] Double fusion coral protein 04: comprising the coral protein amilCP orange, the spider silk repeat region protein Flag, and the coral fluorescent protein Yukon OFP connected in sequence, named amilCP orange-Flag-Yukon OFP, and its amino acid sequence is shown in SEQ ID NO. 15: MSVIAKQMTYKVYMSGTVNGHYFEVEGDGKGKPYEGEQTVKLTVTKGGPLPFAWDILSPQVGYGSIPFTKYPEDIPDYVKQSFPEGYTWERIMNFEDGAVCTVSNDSSIQGNCFIYHVKFSGLNFPPNGPVMQKKTQGWEPNTE RLFARDGMLLGNNFMALKLEGGGHYLCEFKTTYKAKKPVKMPGYHYVDRKLDVTNHNKDYTSVEQCEISIARKPVV AGSSSGSSSGNSVTVSSTVSVGGAGGPGAGGVGLGGAGAGGVGPGGFGGPGGFGGRGGPGGPGGPGGAGGGAGGAG GLYGPGGLYGPGGLYGPGGAGVPGAPGAPGAPGRPGGIGGGAGAGGVGPGGLSGGVGGSGVSVTESVTVSGS MASLSKQVLPRDVRMRFHMDGCVNGHQFTIEGEGAGKPYEGKKTLKLRVTKGGPLPFAFDILSATFTYGNRCFCDYPEDMPDYFKQSLPEGYSWERTLMFEDGGCGTASAHISLEKDCFIHNSTFHGVNFPANGPVMQKKTLNWEPSSELITACDGILKGDVTMFLLLEGGHRLKCQFTTSYKAHKAVKMPPNHIIEHVLVKKEVADGFQIQEHAVAKHFTVDVKET HHHH HH* In the above sequence, the single underlined is amilCP orange, the wavy underlined is the inter-regional linker, the double underlined is the spider silk repeat region protein Flag, the non-underlined is the coral fluorescent protein Yukon OFP, and the dotted underlined is the His tag.

[0069] The DNA sequence of amilCP orange-Flag-Yukon OFP is SEQ ID NO. 16: ATGTCTGTAATCGCAAAACAAATGACCTACAAAGTTTACATGTCTGGTACGGTTAACGGTCATTACTTC GAAGTCGAAGGCGATGGCAAAGGCAAACCGTATGAAGGTGAACAGACCGTGAAACTGACTGTAACCAAAGGTGGTCC TCTGCCATTTGCGTGGGATATTCTGTCTCCGCAGGTTGGATACGGTTCCATCCCGTTTACTAAATACCCGGAAGACA TCCCGGACTACGTAAAGCAGTCTTTTCCGGAGGGCTATACCTGGGAACGCATTATGAACTTCGAAGATGGCGCTGTG TGCACTGTGTCCAATGATTCCTCCATCCAGGGCAACTGTTTTATCTACCACGTTAAATTCAGCGGTCTGAATTTCCC GCCGAACGGTCCGGTCATGCAGAAAAAAACTCAGGGCTGGGAACCGAACACTGAACGTCTGTTCGCTCGTGATGGCA TGCTGCTGGGCAATAACTTCATGGCCCTGAAGCTGGAGGGTGGTGGTCACTACCTGTGTGAGTTCAAAACTACCTAC AAAGCGAAAAAGCCAGTGAAAATGCCAGGTTATCACTATGTGGATCGTAAACTGGATGTGACGAACCACAACAAAGA CTACACTAGCGTTGAACAGTGCGAAATCAGCATCGCTCGTAAGCCGGTAGTGGCGGGCTCAAGCTCCGGAAGTAGCA GTGGTAATAGTGTAACTGTGTCGTCGACGGTTAGTGTCGGAGGAGCGGGCGGTCCCGGAGCAGGCGGTGTCGGTCTA GGTGGTGCGGGTGCAGGCGGCGTAGGTCCAGGTGGTTTCGGCGGACCGGGCGGCTTTGGCGGTCGTGGTGGCCCGGG CGGTCCAGGTGGTCCGGGTGGTGCCGGTGGTGGTGCCGGTGGCGCAGGTGGTCTGTATGGTCCAGGCGGTTTGTACG GGCCGGGGGGACTTTATGGCCCGGGCGGTGCTGGCGTGCCAGGTGCCCCGGGTGCCCCTGGTGCACCTGGCCGCCCT GGCGGCATTGGCGGCGGCGCCGGAGCGGGCGGCGTTGGCCCAGGTGGATTATCTGGTGGTGTCGGCGGCTCAGGTGT CTCGGTTACCGAAAGCGTTACCGTGTCTGGATCCATGGCTTCCCTGTCAAAACAAGTTTTGCCAAGAGATGTCAGAATGCGATTCCACATGGACGGTTGCGTTAATGGTCATCAATTTACCATTGAAGGTGAAGGTGCAGGCAAACCTTACGAGGGTAAAAAAACCCTGAAACTCAGAGTCACAAAGGGGGGTCCTCTCCCTTTTGCCTTTGATATACTGAGTGCTACCTTCACCTATGGCAACCGATGTTTTTGCGATTACCCCGAAGATATGCCAGATTACTTTAAACAAAGCCTGCCCGAGGGCTATTCTTGGGAGCGTACATTAATGTTCGAGGATGGCGGCTGCGGGACCGCCTCTGCTCACATAAGTCTGGAAAAAGATTGTTTTATCCACAATTCCACCTTTCACGGAGTCAACTTCCCAGCTAATGGACCTGTAATGCAGAAGAAGACACTTAACTGGGAACCCAGTTCAGAACTTATCACTGCTTGTGATGGGATACTGAAGGGGGACGTTACCATGTTCTTATTATTGGAAGGAGGACATAGACTTAAATGCCAATTTACAACATCATACAAGGCCCACAAGGCCGTAAAGATGCCCCCCAACCACATTATTGAACACGTCCTGGTGAAGAAAGAAGTAGCTGATGGATTTCAAATCCAGGAACACGCAGTTGCTAAACACTTCACAGTCGATGTCAAGGAAACA CATCATCATCACCATCATTAA In the above sequence, the single underline is the coral protein amilCP orange, the wavy underline is the inter-regional linker, the double underline is the spider silk repeat region protein Flag, the non-underlined is the coral fluorescent protein Yukon OFP, and the dotted underline is the His tag.

[0070] Double fusion coral protein 05: comprising coral protein amilCP, lichen repeat region protein EV1, and coral fluorescent protein mTagBFP2 connected in turn, named amilCP-EV1-mTagBFP2, and its amino acid sequence is SEQ ID NO. 17: MSVIAKQMTYKVYMSGTVNGHYFEVEGDGKGKPYEGEQTVKLTVTKGGPLPFAWDILSPQCQYGSIPF TKYPEDIPDYVKQSFPEGYTWERIMNFEDGAVCTVSNDSSIQGNCFIYHVKFSGLNFPPNGPVMQKKTQGWEPNTE RLFARDGMLLGNNFMALKLEGGGHYLCEFKTTYKAKKPVKMPGYHYVDRKLDVTNHNKDYTSVEQCEISIARKPVV AGSSSGSSSGSGSAAAAAAAAAEAAAAAAAAAAAAGSGAGAGGAGGYGAGAGAGAGAGAGGAAGAGGAGGAGGAGGYGGASVVYVGGGGAGAGAGSGAGAGSGAGAGAGSGAGAGGAGAAAGAGAGAGSGAGSGSGAGAGAGSGAGAGSGAG AGAGAGSGAAGGAGAGAGAGAGGSSGS MVSKGEELIKENMHMKLYMEGTVDNHHFKCTSEGEGKPYEGTQTMRIKVVEGGPLPFAFDILATSFLYGSKTFINHTQGIPDFFKQSFPEGFTWERVTTYEDGGVLTATQDTSLQDGCLIYNVKIRGVNFTSNGPVMQKKTLGWEAFTETLYPADGGLEGRNDMALKLVGGSHLIANAKTTYRSKKPAKNLKMPGVYYVDYRLERIKEANNETYVEQHEVAVARYCDLPSKLGHKLN HHHHHH* In the above sequence, the single underline is amilCP, the wavy underline is the inter-region linker, the double underline is EV1, the non-underlined is mTagBFP2, and the dotted underline is His tag.

[0071] The DNA sequence of amilCP-EV1-mTagBFP2 is SEQ ID NO. 18: ATGTCTGTAATCGCAAAACAAATGACCTACAAAGTTTACATGTCTGGTACGGTTAACGGTCATTACTTC GAAGTCGAAGGCGATGGCAAAGGCAAACCGTATGAAGGTGAACAGACCGTGAAACTGACTGTAACCAAAGGTGGTCC TCTGCCATTTGCGTGGGATATTCTGTCTCCGCAGTGCCAGTACGGTTCCATCCCGTTTACTAAATACCCGGAAGACA TCCCGGACTACGTAAAGCAGTCTTTTCCGGAGGGCTATACCTGGGAACGCATTATGAACTTCGAAGATGGCGCTGTG TGCACTGTGTCCAATGATTCCTCCATCCAGGGCAACTGTTTTATCTACCACGTTAAATTCAGCGGTCTGAATTTCCC GCCGAACGGTCCGGTCATGCAGAAAAAAACTCAGGGCTGGGAACCGAACACTGAACGTCTGTTCGCTCGTGATGGCA TGCTGCTGGGCAATAACTTCATGGCCCTGAAGCTGGAGGGTGGTGGTCACTACCTGTGTGAGTTCAAAACTACCTAC AAAGCGAAAAAGCCAGTGAAAATGCCAGGTTATCACTATGTGGATCGTAAACTGGATGTGACGAACCACAACAAAGA CTACACTAGCGTTGAACAGTGCGAAATCAGCATCGCTCGTAAGCCGGTAGTGGCGGGCTCAAGCTCCGGAAGTAGCA GTGGCTCCGGCTCCGCTGCTGCTGCGGCTGCTGCGGCTGCTGAAGCTGCTGCTGCTGCGGCAGCTGCTGCTGCTGCT GCTGGTTCTGGTGCAGGTGCCGGTGGTGCAGGTGGTTATGGTGCTGGTGCTGGTGCAGGCGCTGGTGCTGGTGCTGG TGGTGCAGCAGGTGCAGGTGGTGCAGGCGGTGCTGGTGGTGCTGGTGGTTACGGCGGCGCAAGCGTAGTATACGTTG GTGGCGGTGGTGCAGGTGCTGGTGCTGGTTCTGGTGCTGGTGCAGGTTCTGGTGCGGGTGCCGGTGCAGGTTCTGGT GCAGGTGCAGGTGGTGCTGGTGCAGCAGCTGGTGCGGGTGCAGGTGCAGGTTCTGGTGCCGGTTCTGGTTCCGGTGC TGGTGCAGGTGCTGGCTCCGGTGCTGGTGCTGGTAGCGGTGCAGGTGCAGGTGCTGGCGCAGGTTCTGGTGCAGCAG GTGGTGCTGGCGCAGGTGCAGGTGCTGGTGCAGGTGGATCCTCTGGATCCATGGTAAGCAAAGGCGAAGAACTGATCAAAGAAAACATGCACATGAAGCTGTACATGGAAGGTACCGTGGATAACCACCATTTCAAGTGCACCTCCGAAGGCGAAGGTAAACCGTACGAGGGTACCCAGACGATGCGCATTAAAGTGGTAGAGGGTGGTCCACTGCCATTTGCATTCGACATTCTGGCTACCTCTTTTCTGTACGGCAGCAAAACCTTCATCAACCACACCCAGGGCATTCCGGACTTTTTTAAGCAGAGCTTCCCTGAAGGCTTTACTTGGGAACGTGTGACTACGTATGAAGACGGCGGCGTTCTGACTGCCACTCAGGACACCTCCCTGCAAGATGGTTGCCTGATCTACAACGTGAAAATCCGCGGTGTCAACTTCACCTCCAACGGTCCGGTTATGCAGAAAAAGACCCTGGGTTGGGAGGCGTTCACCGAGACTCTGTACCCGGCTGACGGCGGTCTGGAGGGTCGCAACGATATGGCTCTGAAACTGGTGGGTGGCTCTCACCTGATTGCGAACGCAAAAACTACGTACCGCAGCAAAAAACCGGCGAAAAACCTGAAAATGCCGGGCGTTTACTACGTGGACTATCGTCTGGAACGTATCAAGGAAGCGAACAACGAAACCTACGTCGAGCAGCATGAAGTAGCTGTAGCACGTTACTGCGATCTGCCGAGCAAACTGGGCCACAAACTGAAC CATCATCAT CACCATCATTAA In the above sequence, the single underlined is the coral protein amilCP, the wavy underlined is the inter-regional linker, the double underlined is the bagworm repeat region protein EV1, the non-underlined is the coral fluorescent protein mTagBFP2, and the dotted underlined is the His tag.

[0072] The above sequences are biosynthesized by silicon, and the above synthesis sequences are inserted into a pET28a commercial plasmid vector, the insertion site is between BamHI and XhoI, and finally the plasmid is transformed into Escherichia coli BL21 (DE3) strain by chemical transformation method (completed by silicon biosynthesis service).

[0073] Example 4 Expression and purification of recombinant double-terminal fusion coral egg protein The strain constructed in Example 3 was fermented, purified, and freeze-dried to obtain a dry powder of the recombinant coral egg protein. The specific implementation is the same as that of Example 2.

[0074] Example 5 Preparation of seaweed fibers by recombinant coral spider silk fusion protein This example relates to a method for preparing calcium alginate fibers of a recombinant coral spider silk fusion protein, and the specific preparation method is as follows: (1) Preparation of protein solution: The freeze-dried powder of the recombinant coral spider silk fusion protein was directly dissolved in ddH2O. After dissolution, the insoluble part was removed by centrifugation at 10,000 rpm for 5 min, and the mass fraction of the protein was controlled to be 1.5%.

[0075] (2) Preparation of recombinant coral spider silk fusion protein-sodium alginate spinning solution: 6 g of sodium alginate was added to 94 g of protein solution, and the mass fraction of sodium alginate was 6 wt%, with a total mass of 100 g. Stir at room temperature for 2-3 h until there are no obvious insoluble substances. After standing for half an hour to remove bubbles, centrifuge at 1000 rpm for 10 min to remove bubbles, and then stand in a 4°C refrigerator overnight to remove bubbles. The obtained spinning solution is used for spinning.

[0076] (3) Preparation of recombinant coral spider silk fusion protein-sodium alginate fibers: The above spinning solution was added to the spinning tube, and wet spinning was carried out using a 0.25 mm pore size spinning needle. The first coagulation bath was a 5 wt% calcium chloride solution, and the second coagulation bath was a 10 wt% calcium chloride solution. The bath temperature was kept at room temperature, the spinning speed was 10 m / min, and the draw ratio was 1:1.2. The spinning line was vertically drawn into the coagulation bath to solidify and form the primary fiber. After washing, oiling, and room temperature drying, the recombinant coral spider silk protein composite seaweed fiber was obtained. The fiber is shown in Figure 4 As shown in the figure (in the figure: white light is natural light, and the conditions for ultraviolet light are: a jade identification strong light flashlight is used as the light source, and the specific flashlight brand is Yedufu, and the article number is SJ-002. The flashlight contains 6 kinds of light sources, which are white light, yellow light, red light, blue light, 365 purple light, and 395 purple light. According to the fluorescence excitation light of different fibers, the light source of the flashlight is selected as follows: single fusion coral protein 01 uses 365 purple light source; double fusion coral protein 01 uses yellow light source; double fusion coral protein 02 uses 365 purple light source; double fusion protein 03 uses 365 purple light source; and double fusion coral protein 05 uses 395 purple light source.), it can be seen that by adding different recombinant coral spider silk proteins, the seaweed fibers present different bright colors. It is proved that the recombinant coral spider silk protein of the present application has a binding force with sodium alginate, and can well composite pigments on the seaweed fibers, providing bright colors for seaweed fibers that are difficult to dye.

[0077] The obtained recombinant coral spider silk fusion protein composite seaweed fiber was subjected to mechanical test, and the dry breaking strength, dry breaking elongation and other properties were tested according to GB / T14337-2022. The sensor reading of the tensile testing machine had hysteresis, and the highest breaking point reading on the machine was taken as the main reading.

[0078] Comparative Example 1 The difference between this comparative example and Example 5 is only that no recombinant coral spider silk fusion protein is added in the process of preparing the recombinant coral composite seaweed fiber, and the other preparation process and test method are exactly the same as those of Example 5.

[0079] Table 1 Mechanical data of Example 5 and Comparative Example 1

[0080] The mechanical data of Comparative Example 1 and Example 5 are shown in Table 1, and the mechanical tensile curve diagram is shown in Figure 5 Compared with pure seaweed fiber, the seaweed fiber added with four different single fusion coral proteins has certain performance improvement, among which the seaweed fiber added with single fusion coral protein 03 has the most obvious performance improvement, the fiber strength is increased by 15.46%, and the elongation is increased by 74.51%. It shows that the recombinant coral spider silk fusion protein of the present application forms a binding force with sodium alginate, and under the action of different recombinant coral spider silk fusion proteins, there are different mechanical property changes, and the mechanical properties of seaweed fiber can be improved to a certain extent.

[0081] Comparative Example 2 The difference between this comparative example and Example 5 is only that no recombinant coral spider silk fusion protein is added in the process of preparing the recombinant coral composite seaweed fiber, and the other preparation process and test method are exactly the same as those of Example 5.

[0082] Table 2 Mechanical data of Example 5 and Comparative Example 2

[0083] The mechanical data of Comparative Example 1 and Example 5 are shown in Table 1, and the mechanical tensile curve diagram is shown in Figure 6As shown, compared with pure seaweed fiber, the seaweed fiber added with 5 different double fusion coral proteins respectively has certain performance improvement, wherein the seaweed fiber added with double fusion coral protein 04 has the most obvious strength performance improvement, and the strength is improved by 20.59%; the seaweed fiber added with double fusion coral protein 05 has the most obvious elongation performance improvement, and the elongation rate is improved by 85.26%. It is shown that the combination between the recombinant coral spider silk fusion protein and sodium alginate is formed, under the action of different recombinant coral spider silk fusion proteins, different mechanical property changes are formed, and the mechanical property of the seaweed fiber can be improved to a certain extent.

[0084] Obviously, the above examples are only examples for clearly illustrating but not limiting the embodiments. Based on the above description, other different forms of changes or variations can be made by those skilled in the art. Here, all the embodiments need not and cannot be exhausted. The obvious changes or variations derived therefrom are still within the protection scope of the present application.

Claims

1. A recombinant coral-spider silk fusion protein, characterized in that, This includes fusing spider silk protein or spider silk protein with coral protein to form recombinant coral-spider silk fusion protein.

2. The recombinant coral-spider silk fusion protein according to claim 1, characterized in that, The spider silk protein or spider silk-like protein includes repeating regions of spider silk protein or bagworm protein.

3. The recombinant coral-spider silk fusion protein according to claim 2, characterized in that, The N-terminus of the spider silk protein or spider silk-like protein also includes spider silk globulin NT; And / or, the C-terminus of the spider silk protein or spider silk-like protein may also include spider silk globulin CT.

4. The recombinant coral-spider silk fusion protein according to claim 3, characterized in that, The spider silk globulin is selected from the NT or CT end of natural spider silk; And / or, the repeating region of the spider silk is selected from the repeating region of any kind of spider silk protein, including but not limited to at least one of MaSp1, MaSp4, Flag, DmarAmSp, MiSp, AcSp, PySp or TuSp; the repeating region of the bagworm protein includes but is not limited to Ev1; Optionally, the amino acid sequence of MaSp1 is as shown in SEQ ID NO.19 or has at least 85%, 88%, 90%, 93%, 95%, 96%, 97%, 98%, 99%, 99.9%, 99.99% or more, or 100% identity with it; its nucleotide sequence is as shown in SEQ ID NO.20 or has at least 85%, 88%, 90%, 93%, 95%, 96%, 97%, 98%, 99%, 99.9%, 99.99% or more, or 100% identity with it. Optionally, the amino acid sequence of MaSp4 is as shown in SEQ ID NO.21 or has at least 85%, 88%, 90%, 93%, 95%, 96%, 97%, 98%, 99%, 99.9%, 99.99% or more, or 100% identity with it; its nucleotide sequence is as shown in SEQ ID NO.22 or has at least 85%, 88%, 90%, 93%, 95%, 96%, 97%, 98%, 99%, 99.9%, 99.99% or more, or 100% identity with it. Optionally, the amino acid sequence of DmarAmSp is as shown in SEQ ID NO.23 or has at least 85%, 88%, 90%, 93%, 95%, 96%, 97%, 98%, 99%, 99.9%, 99.99% or more, or 100% identity with it; its nucleotide sequence is as shown in SEQ ID NO.14 or has at least 85%, 88%, 90%, 93%, 95%, 96%, 97%, 98%, 99%, 99.9%, 99.99% or more, or 100% identity with it. Optionally, the amino acid sequence of the Flag is as shown in SEQ ID NO.25 or has at least 85%, 88%, 90%, 93%, 95%, 96%, 97%, 98%, 99%, 99.9%, 99.99% or more or 100% identity with it; its nucleotide sequence is as shown in SEQ ID NO.26 or has at least 85%, 88%, 90%, 93%, 95%, 96%, 97%, 98%, 99%, 99.9%, 99.99% or more or 100% identity with it. Optionally, the amino acid sequence of Ev1 is as shown in SEQ ID NO.27 or has at least 85%, 88%, 90%, 93%, 95%, 96%, 97%, 98%, 99%, 99.9%, 99.99% or more, or 100% identity with it; its nucleotide sequence is as shown in SEQ ID NO.28 or has at least 85%, 88%, 90%, 93%, 95%, 96%, 97%, 98%, 99%, 99.9%, 99.99% or more, or 100% identity with it. Optionally, the amino acid sequence of the globulin NT is as shown in SEQ ID NO.29 or has at least 85%, 88%, 90%, 93%, 95%, 96%, 97%, 98%, 99%, 99.9%, 99.99% or more, or 100% identity with it; its nucleotide sequence is as shown in SEQ ID NO.30 or has at least 85%, 88%, 90%, 93%, 95%, 96%, 97%, 98%, 99%, 99.9%, 99.99% or more, or 100% identity with it. Optionally, the amino acid sequence of the globulin CT is as shown in SEQ ID NO.31 or has at least 85%, 88%, 90%, 93%, 95%, 96%, 97%, 98%, 99%, 99.9%, 99.99% or more or 100% identity with it; and its nucleotide sequence is as shown in SEQ ID NO.32 or has at least 85%, 88%, 90%, 93%, 95%, 96%, 97%, 98%, 99%, 99.9%, 99.99% or more or 100% identity with it.

5. The recombinant coral-spider silk fusion protein according to any one of claims 1-4, characterized in that, Coral proteins are selected from coral fluorescent proteins and / or coral chromoproteins; Optionally, the coral fluorescent protein includes, but is not limited to, at least one of the following: coral fluorescent protein sfGFP, coral fluorescent protein eforRed, coral fluorescent protein amilCP, coral fluorescent protein mScarlet3, coral fluorescent protein FwYellow, coral fluorescent protein YukonOFP, and coral fluorescent protein mTagBFP2. Optionally, the coral pigments include, but are not limited to, at least one of the following: coral pigments spisPink, coral pigments gfasPurple, coral pigments amilCP orange, and coral pigments amilCP. Optionally, the amino acid sequence of the coral fluorescent protein sfGFP is as shown in positions 350-587 of SEQ ID NO.1 or has at least 85%, 88%, 90%, 93%, 95%, 96%, 97%, 98%, 99%, 99.9%, 99.99% or more, or 100% identity with it; its nucleotide sequence is as shown in positions 1048-1761 of SEQ ID NO.2 or has at least 85%, 88%, 90%, 93%, 95%, 96%, 97%, 98%, 99%, 99.9%, 99.99% or more, or 100% identity with it. Optionally, the amino acid sequence of the coral fluorescent protein eforRed is as shown in positions 227-453 of SEQ ID NO.5 or has at least 85%, 88%, 90%, 93%, 95%, 96%, 97%, 98%, 99%, 99.9%, 99.99% or more, or 100% identity with it; its nucleotide sequence is as shown in positions 679-1359 of SEQ ID NO.6 or has at least 85%, 88%, 90%, 93%, 95%, 96%, 97%, 98%, 99%, 99.9%, 99.99% or more, or 100% identity with it. Optionally, the coral fluorescent protein amilCP is as shown in positions 9-229 of SEQ ID NO.7 or has at least 85%, 88%, 90%, 93%, 95%, 96%, 97%, 98%, 99%, 99.9%, 99.99% or more, or 100% identity with it; its nucleotide sequence is as shown in positions 25-687 of SEQ ID NO.8 or has at least 85%, 88%, 90%, 93%, 95%, 96%, 97%, 98%, 99%, 99.9%, 99.99% or more, or 100% identity with it. Optionally, the coral fluorescent protein mScarlet3 is as shown in positions 317-545 of SEQ ID NO.9 or has at least 85%, 88%, 90%, 93%, 95%, 96%, 97%, 98%, 99%, 99.9%, 99.99% or more, or 100% identity with it; its nucleotide sequence is as shown in positions 949-1635 of SEQ ID NO.10 or has at least 85%, 88%, 90%, 93%, 95%, 96%, 97%, 98%, 99%, 99.9%, 99.99% or more, or 100% identity with it. Optionally, the coral fluorescent protein FwYellow is as shown in positions 342-577 of SEQ ID NO.13 or has at least 85%, 88%, 90%, 93%, 95%, 96%, 97%, 98%, 99%, 99.9%, 99.99% or more, or 100% identity with it; its nucleotide sequence is as shown in positions 1024-1731 of SEQ ID NO.14 or has at least 85%, 88%, 90%, 93%, 95%, 96%, 97%, 98%, 99%, 99.9%, 99.99% or more, or 100% identity with it. Optionally, the coral fluorescent protein YukonOFP is as shown in positions 369-595 of SEQ ID NO.15 or has at least 85%, 88%, 90%, 93%, 95%, 96%, 97%, 98%, 99%, 99.9%, 99.99% or more, or 100% identity with it; its nucleotide sequence is as shown in positions 1105-1785 of SEQ ID NO.16 or has at least 85%, 88%, 90%, 93%, 95%, 96%, 97%, 98%, 99%, 99.9%, 99.99% or more, or 100% identity with it. Optionally, the coral fluorescent protein mTagBFP2 is as shown in positions 400-636 of SEQ ID NO.17 or has at least 85%, 88%, 90%, 93%, 95%, 96%, 97%, 98%, 99%, 99.9%, 99.99% or more, or 100% identity with it; its nucleotide sequence is as shown in positions 1198-1908 of SEQ ID NO.18 or has at least 85%, 88%, 90%, 93%, 95%, 96%, 97%, 98%, 99%, 99.9%, 99.99% or more, or 100% identity with it. Optionally, the coral-colored protein spisPink is as shown in positions 1-224 of SEQ ID NO.9 or has at least 85%, 88%, 90%, 93%, 95%, 96%, 97%, 98%, 99%, 99.9%, 99.99% or more, or 100% identity with it; its nucleotide sequence is as shown in positions 1-672 of SEQ ID NO.10 or has at least 85%, 88%, 90%, 93%, 95%, 96%, 97%, 98%, 99%, 99.9%, 99.99% or more, or 100% identity with it. Optionally, the coral-colored protein gfasPurple is as shown in positions 1-221 of SEQ ID NO.11 or has at least 85%, 88%, 90%, 93%, 95%, 96%, 97%, 98%, 99%, 99.9%, 99.99% or more, or 100% identity with it; its nucleotide sequence is as shown in positions 1-663 of SEQ ID NO.12 or has at least 85%, 88%, 90%, 93%, 95%, 96%, 97%, 98%, 99%, 99.9%, 99.99% or more, or 100% identity with it. Optionally, the coral-colored protein amilCP orange is as shown in positions 1-221 of SEQ ID NO.13 or has at least 85%, 88%, 90%, 93%, 95%, 96%, 97%, 98%, 99%, 99.9%, 99.99% or more, or 100% identity with it; its nucleotide sequence is as shown in positions 1-663 of SEQ ID NO.14 or has at least 85%, 88%, 90%, 93%, 95%, 96%, 97%, 98%, 99%, 99.9%, 99.99% or more, or 100% identity with it. Optionally, the coral-colored protein amilCP is as shown in positions 1 to 221 of SEQ ID NO.17 or has at least 85%, 88%, 90%, 93%, 95%, 96%, 97%, 98%, 99%, 99.9%, 99.99% or more, or 100% identity with it; its nucleotide sequence is as shown in positions 1 to 663 of SEQ ID NO.18 or has at least 85%, 88%, 90%, 93%, 95%, 96%, 97%, 98%, 99%, 99.9%, 99.99% or more, or 100% identity with it.

6. The recombinant coral-spider silk fusion protein according to any one of claims 1-5, characterized in that, The coral protein is fused to at least one end of the spider silk protein or spider silk-like protein.

7. The recombinant coral-spider silk fusion protein according to any one of claims 1-6, characterized in that, The recombinant coral-spider silk fusion protein has a structure from the N-terminus to the C-terminus selected from coral protein-NT globulin-repetitive region-CT globulin, NT globulin-repetitive region-CT globulin-coral protein, coral protein-repetitive region-CT globulin, NT globulin-repetitive region-coral protein, and coral protein A-repetitive region-coral protein B; The coral protein A and coral protein B are respectively selected from coral fluorescent protein and / or coral chromoprotein; Preferably, coral protein A and coral protein B are the same or different.

8. The recombinant coral-spider silk fusion protein according to any one of claims 1-7, characterized in that, A linker peptide is also provided between the spider silk protein or spider silk-like protein and the spider venom protein. And / or, at least one end of the recombinant coral-spider silk fusion protein is also fused with a His tag.

9. The recombinant coral-spider silk fusion protein according to any one of claims 1-7, characterized in that, The recombinant coral-spider silk fusion protein is selected from any one of NT-2Rep-CT-sfGFP, sfGFP-NT-2Rep-CT, NT-2Rep-eforRed, amilCP-2Rep-CT, spisPink-2Rep-mScarlet3, gfasPurple-D3r-mScarlet3, amilCPorange-Dmar-FwYellow, amilCPorange-Flag-YukonOFP, and amilCP-EV1-mTagBFP2.

10. A biomaterial, characterized in that, Includes any one of the following: 1) A nucleic acid molecule encoding the recombinant coral-spider silk fusion protein according to any one of claims 1-9; optionally, the nucleic acid molecule is DNA or RNA; 2) An expression cassette, recombinant vector, recombinant microorganism, or transgenic cell line expressing the nucleic acid molecule of the recombinant coral-spider silk fusion protein according to any one of claims 1-9; 3) Expression cassettes, recombinant vectors, recombinant microorganisms, or transgenic cell lines containing the nucleic acid molecules described in 1); 4) Recombinant vectors, recombinant microorganisms, or transgenic cell lines containing the expression cassettes described in 2) or 3); 5) Host cells containing the recombinant vector described in 2), 3), or 4).

11. The biomaterial according to claim 10, characterized in that, The host cells of the recombinant microorganisms are selected from Escherichia coli of the Enterobacteriaceae family; Optionally, the host cell is selected from Escherichia coli; optionally, the Escherichia coli is Escherichia coli BL21(DE3).

12. A method for preparing the recombinant coral-spider silk fusion protein as described in any one of claims 1-9, characterized in that, include: The recombinant microorganism expressing the recombinant coral-spider silk fusion protein according to any one of claims 1-9 is fermented.

13. A method for preparing functional seaweed fiber, characterized in that, The method includes mixing the recombinant coral-spider silk fusion protein according to any one of claims 1-9 or the recombinant coral-spider silk fusion protein prepared by the preparation method according to claim 12 with a polysaccharide compound to prepare a spinning solution; and using the spinning solution to prepare polysaccharide fibers containing the recombinant coral-spider silk fusion protein.

14. The method for preparing functional seaweed fiber according to claim 13, characterized in that, The spinning solution contains 0.15-1.5% recombinant coral-spider silk fusion protein and 4-10% polysaccharide compounds. And / or, the polysaccharide compound is a polysaccharide compound extracted from seaweed; preferably, the polysaccharide compound extracted from seaweed is selected from at least one of sodium alginate and trehalose.

15. The method for preparing functional seaweed fiber according to claim 13 or 14, characterized in that, Before preparing polysaccharide fibers containing recombinant coral-spider silk fusion protein using the spinning solution, the process further includes a step of degassing the spinning solution.

16. The method for preparing functional seaweed fiber according to any one of claims 13-15, characterized in that, The spinning solution was used to obtain polysaccharide fibers containing recombinant coral-spider silk fusion protein by wet spinning.

17. The method for preparing functional seaweed fiber according to claim 16, characterized in that, The preparation of polysaccharide fibers containing recombinant coral-spider silk fusion protein using the aforementioned spinning solution includes the following steps: adding the spinning solution into a spinning tube, performing wet spinning, and then solidifying the spun fibers in a coagulation bath. Spinning needles with an aperture of 0.1~0.5mm are used; The coagulation bath uses a Ca-containing solution. 2+ Cl - or PO 4- The solution contains ions; preferably, the first coagulation bath is a 5-10 wt% calcium chloride solution, and the second coagulation bath is a 0-10 wt% calcium chloride solution. During spinning, the spinning speed is 10~40m / min; In spinning, the draw ratio is 1:1.1~1.

5.

18. Functional seaweed fiber prepared by a method according to any one of claims 13 to 17.

19. The functional seaweed fiber of claim 18 has any of the following uses: (1) Uses in textiles; (2) In the use of artworks; (3) Used in collectible figures and trendy toys; (4) For use as vehicle decoration; (5) Use of sofas in home use; (6) Used in safety warning items.

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