Synthetic peptides for detection of HIV-1

Abstract

The present invention relates to the field of medical diagnosis, and in particular to a synthetic peptide for detecting HIV-1. The synthetic peptide is obtained by mutating the amino acid sequence shown in SEQ ID NO: 1. It enables better detection of anti-HIV-1 antibodies and better avoids potential "false negative" and "false positive" results.

Description

[0001] This case is a divisional application. The application date of the original application is 2018-08-09, the application number is 2018109041202, and the name of the invention is "Synthetic peptide for detecting HIV-1". Technical Field

[0002] The present invention relates to the field of medical diagnosis, in particular to a synthetic peptide for detecting HIV-1. Background Art

[0003] AIDS, caused by human immunodeficiency virus (HIV), has spread around the world since it was discovered in the 1980s. According to a report by the World Health Organization, by the end of 2016, there were approximately 36.7 million people infected with HIV worldwide. HIV mainly attacks the body's helper T lymphocyte system, selectively invading cells with CD4 molecules, destroying the body's immune capacity, causing the immune system to lose resistance, and often combined with serious opportunistic infections, which leads to various diseases and cancers surviving in the human body, and eventually developing into AIDS. So far, there are no effective drugs or vaccines to treat and prevent AIDS, so HIV infection has become the most serious infectious disease and public health problem threatening the world today.

[0004] HIV virus is an RNA retrovirus with a lipid envelope, and its single-copy gene RNA is about 9.2-9.8Kb in length. HIV has three main structural genes: env, gag, and pol, which encode proteins with different functions, namely antigens. After HIV invades the body, antibodies are generally produced in about 6 weeks. The first to appear in the serum are HIV precursor P55 antibodies and core protein P24 antibodies; then the outer envelope protein GP120 antibodies and transmembrane glycoprotein gp41 antibodies appear. Currently, HIV has been found to be mainly HIV-1 and HIV-2. There are 4 known HIV strains, namely M, N, O, and P. Among them, M and N are the most widely spread. So far, there have been only two P-type cases in the world, and there are only 100,000 O-type cases, mainly concentrated in Central and West Africa. These types are very similar in biological characteristics, and their genomes have 40%-50% homology. At present, the main HIV prevalent in my country is HIV-1.

[0005] Due to the high cost of instruments, reagents and manpower used in HIV antigen and nucleic acid testing, HIV antibody testing still dominates the HIV testing market. At present, HIV antibody testing materials are developing in the direction of improving sensitivity and specificity and shortening the window period on the one hand, and in the direction of simplicity and speed on the other hand. In order to comply with the above development trends, the field also needs to provide HIV-1 recombinant proteins that can be used for HIV-1 antibody testing.

[0006] In view of this, the present invention is proposed. Summary of the invention

[0007] The present invention relates generally to methods and materials for the diagnosis of human immunodeficiency virus (HIV) infection, and more particularly to synthetic peptides having reactive properties binding to HIV-1 type antibodies and their use. The object of the present invention is to provide diagnostic laboratories with a means, in particular a specific peptide, which allows better detection of anti-HIV-1 antibodies and better avoidance of potential "false negative" as well as "false positive" results.

[0008] The synthetic peptide is the amino acid sequence shown in SEQ ID NO:1 including the mutation site;

[0009] The mutation site is selected from at least one of the following:

[0010] T77 was substituted with V, N, or S;

[0011] N101 is replaced by D, E or Q;

[0012] G105 is substituted with K, H or R;

[0013] P124 is replaced by D or E;

[0014] K127 is substituted with D, E or Q;

[0015] Or, an amino acid sequence having at least 80% sequence identity with any of the above sequences, and having a sensitivity greater than 99% and / or a specificity greater than 99.4%. DETAILED DESCRIPTION

[0016] The present invention relates to a synthetic peptide, which is an amino acid sequence shown in SEQ ID NO: 1 including a mutation site;

[0017] The mutation sites are selected from 1, 2, 3, 4 or 5 of the following:

[0018] T77 was substituted with V, N, or S;

[0019] N101 is replaced by D, E or Q;

[0020] G105 is substituted with K, H or R;

[0021] P124 is replaced by D or E;

[0022] K127 is substituted with D, E or Q;

[0023] or, an amino acid sequence having at least 80%, 85%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, 99.5% sequence identity to any of the above sequences;

[0024] In some embodiments, the synthetic peptides, kits or detection methods provided by the present invention have a sensitivity greater than 99%, greater than 99.1%, greater than 99.2%, greater than 99.3%, greater than 99.4%, greater than 99.5%, greater than 99.6%, greater than 99.7%, greater than 99.8%, or greater than 99.9%.

[0025] In some embodiments, the synthetic peptides, kits or detection methods provided by the present invention have a specificity greater than 99.4% (e.g., greater than 99.5%, greater than 99.6%, greater than 99.7%, greater than 99.8%, greater than 99.9% or higher).

[0026] The sensitivity and specificity referred to above are determined according to the invention in the examples of the present invention.

[0027] In some embodiments, the present invention provides a polypeptide comprising all or part of the sequence shown in SEQ ID NO: 1, for example, at least 30 consecutive amino acids.

[0028] The synthetic peptides provided by the present invention are preferably purified and are particularly suitable for the identification and / or enrichment of HIV-1 viruses in animals, especially mammals, especially primates, especially humans. HIV-1 retrovirus is the most common and is dominant in many regions of the world.

[0029] In some embodiments, the mutation sites include: T77 is replaced by V, N or S.

[0030] In some embodiments, the mutation sites include: N101 is replaced by D, E or Q.

[0031] In some embodiments, the mutation sites include: G105 is replaced by K, H or R.

[0032] In some embodiments, the mutation sites include: P124 is replaced by D or E.

[0033] In some embodiments, the mutation sites include: K127 is replaced by D, E or Q.

[0034] In some embodiments, the mutation site further comprises 1, 2, 3, 4 or 5 of the following mutation sites:

[0035] R18 is replaced by S;

[0036] Q95 was replaced by E;

[0037] L106 was replaced by E;

[0038] L123 was replaced by K;

[0039] N130 was substituted with K.

[0040] In some embodiments, the mutation site includes: R18 is replaced by S.

[0041] In some embodiments, the mutation site includes: Q95 is replaced by E.

[0042] In some embodiments, the mutation site includes: L106 is replaced by E.

[0043] In some embodiments, the mutation sites include: L123 is replaced by K.

[0044] In some embodiments, the mutation site includes: N130 is replaced by K.

[0045] In some embodiments, the mutation site is:

[0046] R18 is substituted with S; T77 is substituted with V, N or S; Q95 is substituted with E; N101 is substituted with D, E or Q; G105 is substituted with K, H or R; L106 is substituted with E; L123 is substituted with K; P124 is substituted with D or E; K127 is substituted with D, E or Q; N130 is substituted with K.

[0047] In some embodiments, the mutation site is:

[0048] T77 is substituted with V; Q95 is substituted with E, or not substituted; N101 is substituted with D, E or Q; G105 is substituted with K, H or R; L106 is substituted with E, or not substituted; P124 is substituted with D or EK127 is substituted with D, E or Q.

[0049] In some embodiments, the mutation sites are: R18 is replaced by S, or not replaced; T77 is replaced by V, N or S; Q95 is replaced by E, or not replaced; N101 is replaced by D, E or Q; G105 is replaced by K, H or R; L106 is replaced by E, or not replaced; L123 is replaced by K, or not replaced; P124 is replaced by D or E; K127 is replaced by D, E or Q; N130 is replaced by K, or not replaced.

[0050] In some embodiments, the mutation sites are: R18 is replaced by S; T77 is replaced by S; Q95 is replaced by E, or not replaced; N101 is replaced by D, E or Q; G105 is replaced by K, H or R; L106 is replaced by E, or not replaced; L123 is replaced by K; P124 is replaced by D; K127 is replaced by D, E or Q; N130 is replaced by K.

[0051] In some embodiments, the present invention provides an article comprising the synthetic peptide, such as a microtiter plate (on which the synthetic peptide may be coated).

[0052] The present invention also relates to an isolated nucleic acid molecule encoding a synthetic peptide as described above.

[0053] The present invention also relates to a vector comprising the nucleic acid molecule as described above.

[0054] The present invention also relates to a host cell transformed with the vector described above.

[0055] The host cell may be a eukaryotic cell, such as a mammalian cell.

[0056] The present invention also relates to a method for producing a synthetic peptide as described above, comprising:

[0057] a) expressing a nucleic acid molecule as described above; or

[0058] b) Chemical synthesis, where amino acids are added to obtain the complete polypeptide.

[0059] By the above-mentioned synthesis method a), the expressed polypeptide can be enriched by well-known affinity purification.

[0060] Chemical synthesis can be performed using equipment known to those skilled in the art, such as an automatic peptide synthesizer, such as an automatic peptide synthesizer sold by Applied BioSystems, etc., to obtain the peptide by peptide synthesis.

[0061] The present invention also relates to a modified synthetic peptide obtained by labeling the synthetic peptide as described above with an indicator showing signal intensity.

[0062] In some embodiments, the indicator showing signal intensity includes any one of fluorescent substances, quantum dots, digoxigenin-labeled probes, biotin, radioactive isotopes, radioactive contrast agents, paramagnetic ion fluorescent microspheres, electron-dense substances, chemiluminescent markers, ultrasound contrast agents, photosensitizers, colloidal gold, or enzymes.

[0063] In some embodiments, the fluorescent substance includes Alexa 350, Alexa 405, Alexa 430, Alexa 488, Alexa 555, Alexa 647, AMCA, aminoacridine, BODIPY 630 / 650, BODIPY 650 / 665, BODIPY-FL, BODIPY-R6G, BODIPY-TMR, BODIPY-TRX, 5-carboxy-4′,5′-dichloro-2′,7′-dimethoxyfluorescein, 5-carboxy-2′,4′,5′,7′-tetrachlorofluorescein, 5-carboxyfluorescein, 5-carboxyrhodamine, 6-carboxyrhodamine, 6-carboxytetramethylrhodamine, Cascade Blue, Cy2, Cy3, Cy5, Cy7, 6-FAM, dansyl chloride, fluorescein, HEX, 6-JOE, NBD (7-nitrobenzo-2-oxa-1,3-diazole), Oregon Green 488, Oregon Green 500, Oregon Green514, Pacific Blue, phthalic acid, terephthalic acid, isophthalic acid, cresol fast violet, cresol blue violet, brilliant cresol blue, p-aminobenzoic acid, erythrosine, phthalocyanine, azomethine, cyanine, xanthine, succinylfluorescein, rare earth metal cryptates, trisbipyridyl diamine europium, europium cryptates or chelates, diamine, bicyanine, La Jolla blue dye, allophycocyanin, allococyanin B, phycocyanin C, phycocyanin R, thiamine, phycoerythrin, phycoerythrin R, REG, rhodamine green, rhodamine isothiocyanate, rhodamine red, ROX, TAMRA, Any of TET, TRIT (tetramethylrhodamine isothiol), tetramethylrhodamine, and Texas Red.

[0064] In some embodiments, the radioisotope comprises 110 In, 111 In, 177 Lu, 18 F. 52 Fe, 62 Cu, 64 Cu, 67 Cu, 67 Ga, 68 Ga, 86 Y. 90 Y. 89 Zr, 94 mTc, 94 Tc, 99 mTc, 120 I. 123 I. 124 I. 125 I. 131I. 154-158 Gd, 32 P. 11 C. 13 N. 15 O. 186 Re, 188 Re, 51 Mn, 52 mMn, 55 Co. 72 As, 75 Br, 76 Br, 82 mRb and 83 Any one of Sr.

[0065] In some embodiments, the enzyme includes any one of horseradish peroxidase, alkaline phosphatase, and glucose oxidase.

[0066] In some embodiments, the fluorescent microspheres are polystyrene fluorescent microspheres, which contain rare earth fluorescent ions europium.

[0067] The present invention also relates to a kit comprising the synthetic peptide as described above, or the modified synthetic peptide as described above.

[0068] In some embodiments, the kit further comprises one or more of an immunologically acceptable diluent, a buffer, a protease inhibitor, a blocking agent for blocking non-specific binding of antibodies, and a second antibody having affinity for HIV-1 antibodies.

[0069] In some embodiments, the protease inhibitor can be selected from one or more of PMSF, EDTA, NaN3, pepstantin, leupeptin, aprotinin, Indinavir, Ritonavir, Nelfinavir, Amprenavir, and Kaletra.

[0070] In some embodiments, the second antibody having affinity for HIV-1 antibody can be an antibody against human Fc segment, and the species can be mouse, rat, rabbit, dog, sheep, horse or human;

[0071] In some embodiments, the second antibody having affinity for HIV-1 antibody is labeled with the above-mentioned indicator showing signal intensity.

[0072] In some embodiments, the blocking agent for blocking non-specific binding of antibodies comprises BSA, FBS.

[0073] In some embodiments, the buffer is phosphate buffered saline (PBS), Tris buffered saline (TBS), Tris buffered saline-Tween (TBST) or Tris buffered saline-Triton (TBST).

[0074] In some embodiments, the kit further comprises a microtiter plate, such as described above.

[0075] In some embodiments, when the kit includes a modified synthetic peptide as described above, the kit further includes a reagent for detecting the indicator showing signal intensity;

[0076] The reagents used to detect the indicator showing the signal intensity are well known to those skilled in the art. As an example, when the indicator is horseradish peroxidase (HRP), the corresponding reagents can be selected from hydrogen peroxide and luminol; when the indicator is biotin, the corresponding reagent can be selected from avidin, and so on.

[0077] The present invention also relates to a method for detecting anti-HIV-1 antibodies, which comprises contacting a biological sample with the synthetic peptide as described above, or the modified synthetic peptide as described above, or the reagent in the kit as described above to form an immune complex;

[0078] detecting the presence of the synthetic peptide or the modified synthetic peptide in the immune complex to indicate the presence of HIV-1 antibodies in the biological sample;

[0079] The methods are performed for diagnostic or non-diagnostic purposes, in vivo or in vitro.

[0080] The present invention also relates to the use of the synthetic peptide as described above, or the modified synthetic peptide as described above, or the kit as described above in the preparation of HIV-1 diagnostic agents.

[0081] In some embodiments, provided herein is a method for obtaining or designing a recombinant protein for detecting HIV, which method may include one or more of the following steps:

[0082] Select human immunodeficiency virus envelope protein, select some segments of the protein, and perform mutation design on some sites, prepare the corresponding recombinant protein by expression and purification, and obtain recombinant protein with high activity and good specificity by activity and specificity screening;

[0083] Synthesize sequence fragments and connect them to vectors such as pMD18-T vector to construct T vectors containing the target fragments;

[0084] Through site-directed mutagenesis design, construct a vector containing a specific mutation target fragment, such as pMD18-T vector;

[0085] Preparation method of recombinant protein: design upstream primers (for example, with EcoR I restriction site) and downstream primers (for example, with BamH I restriction site), use a vector containing the target fragment such as T vector as a template, amplify the target gene, double-enzyme digest and purify, then connect to a vector such as pET-28a, and transform cells such as Escherichia coli BL21; after screening to obtain positive clones, pick a single colony and inoculate it into a culture medium such as LB culture medium containing 50ul / ml Kan, and shake culture at an appropriate temperature such as 37 degrees; after OD600 reaches 0.6-0.8, add 1.0mM IPTG, induce culture at 37 degrees for 2-4h, extract total protein, and identify the expression of recombinant protein by SDS-PAGE; use NI ion chelating column to purify the target protein for the first time, use SP column to purify the target protein for the second time, and use SDS-PAGE to identify the purity of the recombinant protein.

[0086] The embodiments of the present invention will be described in detail below in conjunction with the examples, but it will be appreciated by those skilled in the art that the following examples are only used to illustrate the present invention and should not be considered as limiting the scope of the present invention. If no specific conditions are specified in the examples, the conditions are carried out according to normal conditions or the conditions recommended by the manufacturer. If the manufacturer of the reagents or instruments used is not specified, they are all conventional products that can be obtained commercially.

[0087] Example 1

[0088] Design of recombinant proteins

[0089] The gene sequence encoding SEQ ID NO: 1 was synthesized by gene synthesis, and the amino acids at positions X77 (X77 represents the 77th position of the sequence shown in SEQ ID NO: 1, the same below), X95, X101, X105, X106, X124, and X127 were mutated.

[0090] The amino acid at position X77 is mutated from T to V, N or S, the amino acid at position X95 is mutated from Q to E, the amino acid at position X101 is mutated from N to E, D or Q, the amino acid at position X105 is mutated from G to K, H or R, the amino acid at position X106 is mutated from L to E, the amino acid at position X124 is mutated from P to D or E, and the amino acid at position X127 is mutated from K to D, E or Q.

[0091] According to the above mutation direction, mutant clones were constructed, wherein the preferred scheme is that X77, X101, X105, X124, and X127 are respectively V, E, K, D, and D, and are named HIV-Ag-1, respectively N, D, H, E, and E are named HIV-Ag-2, respectively S, Q, R, D, and Q are named HIV-Ag-3, respectively S, D, R, E, and E are named HIV-Ag-4, respectively V, D, K, E, and D are named HIV-Ag-5, respectively N, E, H, D, and E are named HIV-Ag-6, respectively V, Q, R, D, and Q are named HIV-Ag-7, respectively S, Q, K, E, and D are named HIV-Ag-8, and respectively N, Q, H, E, and Q are named HIV-Ag-9. And constructed into pMD18-T vector (TaKara Dalian Bao Biological, catalog number: 6011). The gene sequence of SEQ ID NO: 1 was constructed and named as pMD18-T-HIV-Ag-0, and the mutant clones were named as pMD18-T-HIV-Ag-1 to pMD18-T-HIV-Ag-9 for subsequent amplification and nucleic acid fragment storage.

[0092] Example 2

[0093] Design of recombinant proteins

[0094] Based on Example 1, mutations were designed for the amino acids in the sequences X18, X95, X106, X123 and X130 in HIV-Ag-1 to HIV-Ag-9.

[0095] The amino acid at position X18 was mutated from R to S, the amino acid at position X95 was mutated from Q to E, the amino acid at position X106 was mutated from L to E, the amino acid at position X123 was mutated from L to K, and the amino acid at position X130 was mutated from N to K.

[0096] The new sequences were named HIV-Ag-10 to HIV-Ag-18 and constructed into the pMD18-T vector, named pMD18-T-HIV-Ag-10 to pMD18-T-HIV-Ag-18 for subsequent amplification and nucleic acid fragment storage.

[0097] Example 3

[0098] Construction, induction and purification of recombinant protein expression vector

[0099] Construction and induced expression of recombinant protein expression vector: Design upstream primers (with EcoR I restriction site) and downstream primers (with BamH I restriction site), use pMD18-T-HIV-Ag-0 to pMD18-T-HIV-Ag-18 as templates to amplify the target gene. After the target gene is purified, it is double-digested with EcoR I (TaKara Dalian Bao Biological, Catalog No.: 1010A) and BamH I restriction endonuclease (TaKara Dalian Bao Biological, Catalog No.: 1040A), and incubated at 37°C for 2h. The digested product is purified and connected to the vector pET-28a that has been digested with the same enzymes, incubated at 22°C for 2h and 16°C for 2h. The ligation product was transformed into Escherichia coli BL21 competent (NEB (New England Biolabs), catalog number: C2530H) by heat shock method, and spread on LB plate containing 50ug / ml Kan, and cultured at 37℃ for 16h. Pick the positive clones, identify them by bacterial liquid PCR, and send them for sequencing after double enzyme digestion. Select the positive single clones with completely correct sequencing and inoculate them into LB medium containing 50ug / ml Kan, and culture them at 37 degrees with shaking. After OD600 reaches 0.6-0.8, add 1.0mM IPTG, induce culture at 37℃ for 2-4h, extract total protein, and identify the expression of recombinant protein by SDS-PAGE. Through the 6*HIS tag at the N-terminus of the recombinant protein, the protein with a purity of 96% was obtained after nickel ion chelation purification and SP column purification. The obtained proteins were named HIV-Ag-0 to HIV-Ag-18.

[0100] Example 4

[0101] Application of mutant cloned HIV-Ag-1 to HIV-Ag-9 recombinant antigens in enzyme immunoassay product technology and evaluation of activity and specificity

[0102] The ELISA was performed as follows:

[0103] Coating: Add the recombinant protein to 50mM CB pH 9.6 coating solution at a working concentration of 100ng / ml, mix well and add 100ul per well to the polystyrene plate and coat at 4℃ for 18-20 hours.

[0104] Blocking: Take out the coated plate and equilibrate it at room temperature for 30 minutes, wash the plate twice with washing solution, add 150ul blocking solution to each well and block it at 37℃ for 2 hours. Pat dry and place in an electronic drying oven with humidity less than 30% to dry for 24 hours before use.

[0105] Labeling: The recombinant protein was labeled with HRP according to the recommended method, diluted with enzyme working solution to 50 ng / ml and mixed evenly (enzyme diluent formula: 20 mM PB 150 mM NaCl 0.5% BSA 0.05% Tween-200 0.1% P300).

[0106] Reaction mode and reaction time: 50ul of the sample to be tested + 50ul of the sample diluent, react in a 37-degree thermostat for 60min; wash the plate 5 times, pat dry and add 100ul of the labeled recombinant protein-HRP working solution, react at 37°C for 30min; wash the plate 5 times, add 50ul of color developer A and B respectively, and develop for 30min; add 50ul of the stop solution, use dual wavelength detection at 450nm and 630nm, and the detection must be completed within 10min.

[0107] The performance comparison experiment was conducted with the commercially available kit, with 500 HIV-1 positive sera and 3000 HIV-1 antibody negative sera. The results are shown in Table 1. The recombinant protein of the present invention is used for HIV-1 antibody detection, and the sensitivity is improved and the specificity is significantly improved, which is better than the existing products.

[0108] Table 1 Detection results after mutation

[0109]

[0110] Example 5

[0111] Activity evaluation of mutant cloned HIV-Ag-1 to HIV-Ag-18 recombinant antigens in ELISA process The ELISA test was performed as follows:

[0112] Coating: Add the recombinant protein to 50mM CB pH 9.6 coating solution at a working concentration of 100ng / ml, mix well and add 100ul per well to the polystyrene plate and coat at 4℃ for 18-20 hours.

[0113] Blocking: Take out the coated plate and equilibrate it at room temperature for 30 minutes, wash the plate twice with washing solution, add 150ul blocking solution to each well and block it at 37℃ for 2 hours. Pat dry and place in an electronic drying oven with humidity less than 30% to dry for 24 hours before use.

[0114] Labeling: The recombinant protein was labeled with HRP according to the recommended method, diluted with enzyme working solution to 50 ng / ml and mixed evenly (enzyme diluent formula: 20 mM PB 150 mM NaCl 0.5% BSA 0.05% Tween-200 0.1% P300).

[0115] Reaction mode and reaction time: 50ul of the sample to be tested + 50ul of the sample diluent, react in a 37-degree thermostat for 60min; wash the plate 5 times, pat dry and add 100ul of the labeled recombinant protein-HRP working solution, react at 37°C for 30min; wash the plate 5 times, add 50ul of color developer A and B respectively, and develop for 30min; add 50ul of the stop solution, use dual wavelength detection at 450nm and 630nm, and the detection must be completed within 10min.

[0116] 500 HIV-1 positive sera were diluted with sample diluent at 1:200 and used as test specimens for activity detection. The results are shown in Table 2. After mutation, the average sensitivity of the 500 positive specimens after dilution was significantly improved.

[0117] Table 2 Detection results after mutation

[0118]

[0119]

[0120] Finally, it should be noted that the above embodiments are only used to illustrate the technical solutions of the present invention, rather than to limit it. Although the present invention has been described in detail with reference to the aforementioned embodiments, those skilled in the art should understand that they can still modify the technical solutions described in the aforementioned embodiments, or replace some or all of the technical features therein by equivalents. However, these modifications or replacements do not make the essence of the corresponding technical solutions deviate from the scope of the technical solutions of the embodiments of the present invention.

Claims

1. A synthetic peptide, characterized in that The synthetic peptide is obtained by simultaneously performing the following mutations on the basis of the amino acid sequence shown in SEQ ID NO: 1: R18 is substituted with S, Q95 is substituted with E, L106 is substituted with E, L123 is substituted with K, N130 is substituted with K, T77 is substituted with V, N101 is substituted with D, G105 is substituted with K or R, P124 is substituted with E, and K127 is substituted with D or E.

2. A synthetic peptide, characterized in that The synthetic peptide is obtained by simultaneously performing the following mutations on the basis of the amino acid sequence shown in SEQ ID NO: 1: R18, Q95, L106, L123, N130 are replaced by S, E, E, K, K, respectively, and T77, N101, G105, P124, K127 are replaced by any one of the following 9 groups: 。 3. An isolated nucleic acid molecule, characterized in that It encodes the synthetic peptide according to any one of claims 1 to 2.

4. A carrier, characterized in that It comprises the nucleic acid molecule according to claim 3.

5. A host cell, characterized in that It is transformed with the vector described in claim 4.

6. A method for preparing the synthetic peptide according to any one of claims 1 to 2, characterized in that: include: a) expressing the nucleic acid molecule according to claim 3; or b) Chemical synthesis, adding amino acids to obtain a complete polypeptide, said polypeptide being a synthetic peptide as claimed in claim 1 or 2.

7. A modified synthetic peptide, characterized in that The method is obtained by labeling an indicator showing signal intensity with the synthetic peptide according to any one of claims 1 to 2.

8. The modified synthetic peptide according to claim 7, characterized in that The indicator showing signal intensity includes a fluorescent substance.

9. The modified synthetic peptide according to claim 7, characterized in that The indicator showing signal intensity includes any one of quantum dots, digoxigenin-labeled probes, biotin, radioactive isotopes, radioactive contrast agents, paramagnetic ion fluorescent microspheres, electron-dense substances, chemiluminescent markers, ultrasound contrast agents, photosensitizers, colloidal gold or enzymes.

10. A kit, characterized in that: It includes the synthetic peptide according to any one of claims 1 to 2, or the modified synthetic peptide according to any one of claims 7 to 9.

11. The kit according to claim 10, characterized in that The kit may further include one or more of an immunologically acceptable diluent, a buffer, a protease inhibitor, a blocking agent for blocking non-specific binding of the antibody, and a second antibody having affinity for the HIV-1 antibody.

12. The kit according to any one of claims 9 to 11, characterized in that The kit also includes a reagent for detecting the indicator showing signal intensity.

13. A method for detecting anti-HIV-1 antibodies in vitro for non-diagnostic purposes, characterized in that: The method comprises contacting a biological sample with the synthetic peptide according to any one of claims 1 to 2, or the modified synthetic peptide according to any one of claims 7 to 9, or the reagent in the kit according to any one of claims 10 to 12 to form an immune complex; The presence of the synthetic peptide or the modified synthetic peptide in the immune complex is detected to indicate the presence of HIV-1 antibodies in the biological sample.

14. Use of the synthetic peptide according to any one of claims 1 to 2, or the modified synthetic peptide according to any one of claims 7 to 9, or the kit according to any one of claims 10 to 12 in the preparation of an HIV-1 diagnostic agent.

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