DNAzyme capable of specifically recognizing and detecting decapod iridovirus 1 and its application

Abstract

The present invention relates to the technical field of pathogenic microorganism detection, and specifically discloses a DNAzyme capable of specifically identifying and detecting decapod iridescent virus 1 and an application thereof, including a deoxyribonucleotide sequence of DNAzyme MCP-1. The present invention obtains the DNAzyme MCP-1 by screening through the SELEX technology, and adds SYBR Green to amplify the fluorescence signal, so as to prepare a biosensor capable of identifying decapod iridescent virus 1 with high specificity and high sensitivity. The DNAzyme has high thermochemical stability, is easy to synthesize and modify, and has low cost. It is not only convenient to cooperate with signal amplification means such as nucleic acid amplification, but also easy to use various signal transduction mechanisms to be integrated into analysis and reporting systems such as fluorescence.

Description

Technical Field

[0001] The present invention relates to the technical field of pathogenic microorganism detection, in particular to a DNAzyme capable of specifically identifying and detecting decapod iridovirus 1 and an application thereof. Background Art

[0002] In recent years, diseases have frequently occurred in shrimp farming, resulting in a decrease in production and great economic losses. Among them, viral diseases are the main diseases in shrimp farming and one of the main factors affecting the development of shrimp farming.

[0003] Decapod Iridescent Virus 1 (DIV1) belongs to the Iridoviridae family. It is a double-stranded DNA virus with an icosahedral symmetric structure. It spreads rapidly among decapod crustaceans and has a wide host range. It is mainly distributed in coastal shrimp and crab farming areas. The mortality rate of shrimp infected with DIV1 is extremely high. Clinical symptoms include: basophilic inclusion bodies and nuclear condensation in the hepatopancreas and muscle tissues. Shrimp will swim, lie on their sides, have a slightly reddish body and an empty stomach. At present, there is no available drug treatment for DIV1. The management of DIV1 at home and abroad is mainly based on comprehensive prevention, that is, early detection of the disease and taking corresponding measures are important ways to control the spread of the virus.

[0004] Traditional methods for detecting whether shrimp are infected with DIV1 mainly include: pathological microscope observation, biochemical determination, immunological test, cell culture, etc. These methods are time-consuming and complicated to operate, and are difficult to meet the needs of rapid on-site detection.

[0005] Deoxyribozymes (DNAzymes) are isolated through in vitro screening by ligand evolution by exponential enrichment (SELEX) and are usually composed of a substrate chain and an enzyme chain. These two DNA chains partially complement each other through base pairing to form a double-stranded system. The presence of the target activates the activity of the DNA enzyme, and the nucleotide (rA) on the substrate chain is cleaved. The DNAzyme can complete the conduction and amplification of the signal through conformational changes and cascade reactions, so that it can perform highly sensitive detection when used as a biosensor. In addition, DNAzyme also has the advantages of good stability, easy modification and preparation, etc., providing an ideal molecular tool for the development of sensitive and efficient on-site detection methods.

[0006] The present invention uses SELEX technology to screen out DNAzymes that can specifically recognize DIV1, and designs a biosensor based on DNAzyme's specific recognition of DIV1. By optimizing its reaction conditions, a biosensor for detecting DIV1 with high specificity and good sensitivity is obtained, which can quickly detect aquatic pathogenic microorganisms DIV1. Summary of the invention

[0007] The purpose of the present invention is to address the defects of the prior art and provide a DNAzyme capable of specifically identifying and detecting Decapoda iridovirus 1 and its application, so as to solve the problems raised by the above-mentioned background technology.

[0008] To achieve the above object, the present invention provides the following technical solution: a DNAzyme capable of specifically identifying and detecting decapod iridescent virus 1, comprising a deoxyribonucleotide sequence of DNAzyme MCP-1:

[0009] The MCP-1 is: 5'-GAAAAGTGTTATCCGGGGAAAGATCATCGTACTGTT ATCTCCGAGCCGGTCGACCAGCATCGGTAGGCGTATCT-3'.

[0010] As a preferred technical solution of the present invention, the DNAzyme sequence has a labeled quenching group at the 3' end.

[0011] The invention discloses a method for screening DNAzyme capable of specifically recognizing and detecting decapod iridovirus 1, and adopts a magnetic bead SELEX method to screen the DNAzyme.

[0012] As a preferred technical solution of the present invention, the specific steps are as follows:

[0013] Step 1: The library, primers, and plasmids used in the experiment were designed by ourselves and synthesized at Shanghai Bioengineering;

[0014] Step 2: The plasmid containing the capsid protein sequence of decapod iridovirus 1 is transferred into the engineered bacteria, induced to express and purified and recovered, and the same method is used to obtain the reverse screening crude protein from the empty engineered bacteria;

[0015] Step 3: The crude protein obtained in step 2 is tested for crude protein concentration, divided into 1.5 mL sterile EP tubes, and stored at -20°C for future use;

[0016] Step 4: The initial library containing 40 random bases in the central region and 20 fixed bases at both ends is connected with primers through PCR reaction, rA cleavage site and biotin label are introduced, and DNA is recovered by gel cutting;

[0017] Step 5: Connect the PCR product obtained in step 4 to the streptavidin-coated magnetic beads at 30°C for 30 minutes;

[0018] Step 6: After the connection is completed, the supernatant is discarded by magnetic separation, and the magnetic beads are washed three times with avidin reaction buffer;

[0019] Step 7: Wash the magnetic beads twice with 0.2 M NaOH and separate them magnetically to prepare single strands;

[0020] Step 8: Wash the magnetic beads 3 times with ultrapure water to make the pH value close to neutral;

[0021] Step 9: 10 μg of capsid protein of decapod iridovirus 1 (dissolved in 50 mM PBS buffer, pH 8.0) was mixed with an equal volume of 2× screening buffer, incubated at 30° C. for 1 h to allow cleavage reaction to occur, magnetic separation was performed, and the supernatant was recovered by alcohol precipitation;

[0022] Step 10: Using the product obtained in step 9 as a template, add a primer containing biotin for PCR amplification for the next round of screening;

[0023] Step 11: High-throughput sequencing of the PCR products obtained in the ninth round and DNA sequence analysis;

[0024] Step 12: Detection of activity and properties of candidate DNAzymes;

[0025] Step 13: Immobilize the DNAzyme complex in a polyethylene 96-well plate using trehalose and pullulan.

[0026] As a preferred technical solution of the present invention, biotin is labeled at the 5' end of the DNA sequence; DNA is combined with magnetic beads, the cleavage reaction, and alcohol precipitation recovery are all carried out in a 1.5 mL sterile EP tube;

[0027] DNA concentration was measured using an ultra-micro-volume UV spectrophotometer.

[0028] As a preferred technical solution of the present invention, a fluorescence signal detector is used to detect the intensity of DNAzyme cutting fluorescence.

[0029] As a preferred technical solution of the present invention, the diameter of the magnetic beads used in the DNAzyme screening process is 0.5 μm.

[0030] Application of a DNAzyme system that can specifically identify and detect Decapoda iridovirus 1 in aquatic product testing.

[0031] A DNAzyme capable of specifically recognizing and detecting decapod iridovirus 1 is used in the preparation of a biosensor capable of rapidly detecting decapod iridovirus 1.

[0032] A kit capable of specifically identifying and detecting decapod iridovirus 1, the kit containing DNAzyme.

[0033] Compared with the prior art, the beneficial effects of the present invention are as follows: the DNAzyme obtained by screening through the SELEX technology is prepared into a DNAzyme system so that DIV1 can be identified with higher specificity and high sensitivity, and the DNAzyme has high thermochemical stability, is easy to synthesize and modify, and has low cost. It is not only convenient to cooperate with signal amplification means such as nucleic acid amplification, but also easy to use various signal transduction mechanisms to be integrated into analysis and reporting systems such as fluorescence. The DNAzyme-based biosensor of the present invention is used to quickly detect DIV1 of aquatic products, and the method is simple and easy to operate. Therefore, the present invention has good application prospects in the detection of DIV1. BRIEF DESCRIPTION OF THE DRAWINGS

[0034] Figure 1 This is a diagram of the expression and purification of DIV1 capsid protein;

[0035] Figure 2 This is a flow chart for screening DNAzymes according to the present invention;

[0036] Figure 3 This is a secondary structure simulation diagram of the DNAzyme MCP-1 screened in the present invention;

[0037] Figure 4 It is the activity and specificity detection diagram of MCP-1 in the present invention;

[0038] Figure 5 This is a gel image showing the cleavage activity and specificity detection of MCP-1 in the present invention;

[0039] Figure 6 This is a diagram for optimizing the pH value affecting the MCP-1 cleavage reaction in the present invention;

[0040] Figure 7 This is a diagram for detecting metal ions that affect the MCP-1 cleavage reaction in the present invention;

[0041] Figure 8 This is a detection diagram of the biosensor of the present invention for different concentrations of DIV1. DETAILED DESCRIPTION

[0042] The preferred embodiments of the present invention are described in detail below in conjunction with the accompanying drawings so that the advantages and features of the present invention can be more easily understood by those skilled in the art, thereby making a clearer and more definite definition of the protection scope of the present invention.

[0043] Example 1: A method for screening a DNAzyme that can specifically recognize decapod iridovirus 1, the specific steps are as follows:

[0044] Step 1: Synthesize a random single-stranded DNA library and primers for the following sequences:

[0045] Initial random library:

[0046] 5'-GGATAACACTTTTCTGTTAT-N40-CAGCATCGGTAGGCGTATCT-3';

[0047] The forward primer was: 5′-ACCGATGCTGTrAGGATAACACTTTTC-3′;

[0048] The backward primer was: 5′-AGATACGCCTACCGATGCTG-3′;

[0049] Substrate: 5'-FAM-AGATACGCCTACCGATGCTGT / rA / GGATAACACTTTTC-3'. Quencher substrate: 5'-CAGCATCGGTAGGCGTATCT-BHQ1-3'.

[0050] Step 2: Cloning, expression and purification of the major capsid protein of decapod iridovirus 1;

[0051] Step 2.1, primer design and expression vector construction;

[0052] The protein coding sequence of the major capsid protein of decapod iridovirus 1 was obtained from GenBank (accession number: ATE87157.1). Shanghai Bioengineering was commissioned to reverse translate it and optimize its expression in Escherichia coli to synthesize the plasmid PET29a containing the target sequence. Through bioinformatics analysis, primers were designed with BamHⅠ and XhoⅠ as restriction sites: F-BamHⅠ(5'-CGCGGATCCATGCTGCGTTTCATCTACGAAAAGAA-3'); R-XhoⅠ(5'-CCGCTCGAGCATGAACGGCAGGCCCAG-3');

[0053] Step 2.2, expression of the major capsid protein of decapod iridovirus 1;

[0054] Transform the plasmid into E.coli DH5α competent cells, ice bath for 30 minutes, heat shock in 42℃ water bath for 70 seconds, quickly ice bath for 2:30 minutes, add 600μL LB liquid medium without antibiotics, mix well, culture at 37℃, 180rpm for 1-1.5h, take 100μL of bacterial solution and spread it on LBK solid medium containing 30μg / mL kanamycin, and culture at 37℃ for 12-15h; culture at 37℃ for 6-8h with LBK solid medium, pick positive bacteria for colony PCR and sequencing identification; use the kit to extract the plasmid, and use the same method to transform the extracted E.coli DH5α plasmid into E.coli BL21 competent cells;

[0055] Step 2.3, expression of the major capsid protein of decapod iridovirus 1;

[0056] A single colony was inoculated into LBK liquid medium and cultured at 37°C and 180 rpm for 4 to 6 h. 600nm When the inoculum was 0.8, LBK liquid medium was inoculated with 4% inoculum and cultured at 37℃ and 180r / min for 4-5h. 600nm When the pH value is about 0.6 to 0.8, add isopropyl-β-D-thiogalactoside with a final concentration of 1 mmol / L, the temperature is 37°C, and ferment for 4 hours; centrifuge the cultured bacterial solution at 8000r / min for 15 minutes, discard the supernatant, resuspend the precipitated bacteria in PBS buffer, ultrasonically disrupt for 30 minutes, and centrifuge at 8000r / min for 10 minutes. The supernatant is the crude protein solution, and the reverse screening protein is the crude protein solution prepared by the unloaded E.coli BL21 competent cells according to the above steps;

[0057] Step 2.4, purification of the major capsid protein of decapod iridovirus 1;

[0058] After the crude protein solution is loaded onto SDS-PAGE gel electrophoresis, the gel block is taken out and stained with 0.25M KCl precooled at 4°C. A white band will appear at the target band after 1 to 10 minutes. The gel band where the target protein is located is cut out with a sterilized blade, and the gel strip is soaked in distilled water for 5 minutes until it becomes colorless and transparent. The gel strip is taken out from the distilled water and placed in the prepared 1.5mL sterile centrifuge tube, crushed with sterile tweezers, and 1mL PBS is added. The centrifuge tube is placed in a 4°C refrigerator for 24 hours, the liquid in the tube is aspirated, and a single band is detected by SDS-PAGE gel electrophoresis. The purified protein is then packaged and stored in a -20°C refrigerator.

[0059] Step 3: Screening process Figure 1 As shown, the magnetic bead SELEX method was used to screen DNAzyme:

[0060] Step 3.1, magnetic beads bind to the library;

[0061] Take 50 μL of deoxyribonucleotide library and dilute it to 500 μL with Buffer I, mix it with 100 μL of magnetic beads, place it on a rotating mixer, and incubate it at 30°C, 1000 r / min for 30 min. After magnetic separation, discard the supernatant, and wash the magnetic beads three times with Buffer I;

[0062] Step 3.2, removing DNA without biotin (preparing single strands);

[0063] Wash the magnetic beads twice with 500 μL 0.2 mol / L NaOH, place at room temperature for 2 min, perform magnetic separation, and discard the supernatant (wash the magnetic beads several times with deionized water to ensure that the pH value is maintained at around 7.0);

[0064] Step 3.3, negative screening;

[0065] Negative screening was added only in the third and fifth rounds of screening. The magnetic beads were resuspended in 150 μL of 2× screening buffer solution, 20 μg of anti-screening protein was added, and PBS was added to make the final volume reach 300 μL. The beads were placed on a rotating mixer and incubated at 30°C, 800 r / min for 60 min. The beads were separated magnetically and the supernatant was discarded. The beads were washed twice with 500 μL of 2× screening buffer solution, separated magnetically, and the supernatant was discarded.

[0066] Step 3.4, positive screening;

[0067] Take 150 μL of 2× screening buffer solution to resuspend the magnetic beads, then add 20 μg of purified protein, add PBS to make the final volume reach 300 μL, incubate on a rotating mixer at 30°C, 800 r / min for 60 min, magnetically separate, and collect the supernatant;

[0068] Step 3.5, recovering the DNA library by alcohol precipitation;

[0069] Add 0.1 times the volume of 3M NaOAc, mix well, then add 900μL of 100% ice ethanol, mix well, and place at -20℃ for 1h. Centrifuge at 12000g, 4℃ for 20min in a refrigerated centrifuge, and discard the supernatant. Add 200μL of 70% ice ethanol, gently rinse the inside of the EP tube with a pipette tip, centrifuge at 12000g, 4℃ for 10min, and discard the supernatant. Open the EP tube cap and vacuum dry at 30℃ for 30min. Add 100μL of deionized water to dissolve and shake to mix, and you will get a DNAzyme secondary library with cleavage activity against the major capsid protein of Decapoda iridovirus 1.

[0070] 2× Screening Buffer Solution: 100 mM HEPES (pH 7.5), 300 mM NaCl, 30 mM MgCl 2 , 0.02% Tween-20; the Buffer I formula is: 10mM Tris-HCl (pH 7.5), 1mM EDTA, 1M NaCl, 0.01% to 0.1% Tween-20.

[0071] Step 3.6, PCR amplification and library screening;

[0072] The DNAzyme secondary library obtained in step 3.5 was amplified by PCR. The PCR reaction system with a total volume of 50 μL was:

[0073] DNA template (20 ng / μL): 1 μL;

[0074] Forward primer FP (100 μM): 1 μL;

[0075] Backward primer RP (100 μM): 1 μL;

[0076] Taq DNA polymerase (5U / μL): 25μL;

[0077] MgCl 2 :5μL;

[0078] dH 2 O: 17 μL;

[0079] PCR amplification conditions: pre-denaturation at 95°C for 3 min; then 27 cycles of denaturation at 95°C for 15 s; annealing at 56.8°C for 15 s; extension at 72°C for 1 min, and finally extension at 72°C for 5 min.

[0080] The DNA sequences in the PCR products were recovered using a gel recovery kit as a secondary library.

[0081] Step 4: Repeat the screening as above;

[0082] The recovered products of each round of PCR are used as the screening library for the next round, and the SELEX screening in step 3 is repeated 9 times. The DNAzyme secondary library obtained in the last round of screening is the enriched library obtained by screening.

[0083] Step 5: High-throughput sequencing analysis;

[0084] High-throughput sequencing was completed by Shanghai Bioengineering using the Miseq technology of Illumina, and a total of 161,054 original DNA sequences were obtained. After homology analysis of the 20 sequences with the highest enrichment rate, 4 sequences were selected for synthesis for affinity verification, and finally the sequence with the highest affinity, MCP-1, was selected for analysis. The sequence proportions are shown in Table 1.

[0085] Table 1: DNA sequences obtained by high-throughput sequencing and their proportions

[0086]

[0087] Example 2: Preparation of a DNAzyme system that can specifically recognize decapod iridovirus 1 and verification of its cleavage activity and specificity,

[0088] The specific steps include:

[0089] Step 1: Candidate DNA sequence activity verification:

[0090] Step 1.1: Preparation of decapod iridovirus 1 (DIV1) suspension;

[0091] Select white shrimp infected with decapod iris virus 1 (DIV1), remove the cephalothorax tissue from the cephalothorax and cut it into pieces in a beaker, then add 40mL of pre-cooled PPB-Tris (376.07mM sodium chloride, 6.32mM potassium sulfate, 6.4mM magnesium sulfate, 14.41mM calcium chloride, 50mM Tris-HCl, pH6.5-8.0) to the beaker, grind it with a glass grinder, pour the resulting mixture into a 50mL centrifuge tube, centrifuge it at 4℃, 10000rpm for 10min, and collect the supernatant. Repeat the above operation to collect the supernatant. First, filter the collected supernatant through a 500-mesh sieve to remove the coarser material, and then filter it with a 0.45μm filter membrane. After the virus suspension is prepared, take out a portion for multiple pathogen detection, ensure that there is only DIV1 in the virus suspension, and freeze it at -80℃.

[0092] Step 1.2: Fluorescence signal monitoring method;

[0093] In this study, we used a kinetic method to monitor the fluorescence signal to prove that it has a cleavage reaction. The DNAzyme complex is composed of 2μL substrate, 2.6μL MCP-1, 15.4μL PBS, and 20μL 2× screening buffer. After mixing in a light-proof tube, it is boiled in a water bath for 5 minutes and then placed at room temperature for half an hour to fold into a better secondary structure to obtain the DNAzyme complex MCP-1-S. The fluorescent signal sensor is composed of 45μL 2× screening buffer, 5.57*10 6 The samples were composed of 100 μL of PBS, 100 μL of 4 μL of decapod iridovirus 1 (DIV1) and 4 μL of DNAzyme complex. When DIV1 was added, a cleavage reaction occurred, generating a fluorescent signal. The fluorescence signal was continuously monitored for 1 hour using an ELISA reader (Infinite M1000Pro, Tecan, Switzerland) with an excitation wavelength of 486 nm and an emission wavelength of 524 nm, and the detection was performed every 30 seconds. PBS buffer was used as a control. Each experiment was repeated three times to draw a kinetic curve.

[0094] Step 1.3: denaturing polyacrylamide gel electrophoresis (15% dPAGE);

[0095] To further verify the DNAzyme cleavage reaction, 4 μL MCP-1-S, 25 μL 2× Selection buffer and 5.57*10 610 copies DIV1, make up to 50 μL with PBS to prepare the reaction solution, react for 20 min in dark conditions, and terminate the reaction with 2× gel blue dye (containing 8M urea). Take an appropriate amount of sample and load it on 15% dPAGE, and separate it by electrophoresis at a voltage of 120V for 50 min. Use Bio-Rad GelDocTM EZ imaging system (BIO-RAD, USA) to image the gel results and quantitatively analyze the bands (DNA content of the cut band + the uncut band = 100%).

[0096] Step 2: Specificity detection;

[0097] The DNAzyme (MCP-1) system was as follows: 4 μL MCP-1-S, 45 μL 2× screening buffer and 41 μL PBS were mixed evenly, and 5.57*10 6 DIV1 copies were added with water, 100 μg of anti-sieve protein, and the shrimp tissue extract not infected with DIV1 was used as the control group. The fluorescence signal was monitored within 1 hour on a full-wavelength multifunctional microplate reader (excitation wavelength of 486 nm and emission wavelength of 524 nm). Figure 4 and Figure 5 As shown, MCP-1 can only produce a cleavage effect on Decapoda Iridovirus 1 and show a strong fluorescence signal.

[0098] Step 3: Optimization of reaction conditions;

[0099] Step 3.1: pH optimization;

[0100] The appropriate pH environment, cleavage reaction time and concentration play a crucial role in the realization of the sensor. When we optimize the pH value of the buffer in the reaction, the optimization range is pH (4.5-9.0), such as Figure 6 As shown in the figure, the cleavage activity of DNAzyme is the highest at pH 8.0, which is the optimal pH for DNAzyme.

[0101] Step 3.2: Optimization of metal ions;

[0102] Two monovalent metal ions (K + , NH4 + , final concentration of 50 mM, pH adjusted to the optimal value) and 8 divalent metal ions (Ba 2+ , Ca 2+ , Cu 2+ , Fe 2+ , Mn 2+ , Ni 2 , Sr. 2+ , Zn 2+, final concentration of 50 mM, pH adjusted to optimal) to prepare the buffer, incubate with ethylenediaminetetraacetic acid (EDTA) and incubate with Na + It is a monovalent metal ion, Mg 2+ For divalent metal ions, use them as the control group for ion experiments. Take 4 μL MCP-1-S, 45 μL different ion buffers, 41 μL ddHO 2 O and 5.57*10 6 After mixing copiesDIV1 evenly, use a fluorescence microplate reader to monitor the fluorescence response within 1 hour (excitation wavelength 486nm, emission wavelength 524nm, record every 30s). Test the effect of different divalent metal ions on the cleavage activity of MCP-1 and determine the optimal metal ion. Figure 7 As shown in Figure 2, DNAzyme has almost no cleavage activity without the assistance of divalent metal ions (EDTA group). + The fluorescence response of Mg is the strongest among the divalent metal ions. 2+ The fluorescence response of Na + As the best monovalent ion, Mg 2+ As the best divalent ion.

[0103] Example 3: Application of a DNAzyme-based biosensor capable of specifically identifying decapod iridovirus 1 in rapid detection of aquatic products.

[0104] The specific steps are as follows:

[0105] Prepare a polyethylene 96-well plate, fix the MCP-1 complex with trehalose and pullulan to prepare a fluorescent biosensor.

[0106] Take the infected shrimp meat, add physiological saline, grind and centrifuge, and retain the supernatant.

[0107] The sensor system is as follows: 4 μL MCP-1-S, 45 μL 2× screening buffer and 41 μL PBS were mixed, 1 μL SYBR Green was added and mixed to amplify the fluorescence signal, 0.8 μL quenching substrate was added and allowed to stand for 5 min to quench the free fluorescent groups in the system, and then the initial fluorescence signal was monitored on a full-wavelength multifunctional microplate reader (excitation wavelength of 486 nm, emission wavelength of 524 nm). 6 The shrimp tissue extracts not infected with DIV1 were added as the control group, and the fluorescence signals were observed on a full-wavelength multifunctional microplate reader (excitation wavelength of 486 nm, emission wavelength of 524 nm) after 10 min of reaction to calculate the relative fluorescence.

[0108] The supernatant with different amounts of DIV1 virus suspension was used as the experimental group, and the shrimp tissue extract that was not infected with DIV1 was added as the control. The results showed that as the amount of DIV1 in the system increased, the fluorescence signal of the experimental group increased significantly, while the fluorescence signal of the negative control group did not change. Figure 8 shown.

[0109] The above embodiments only express the implementation methods of the present invention, and the descriptions thereof are relatively specific and detailed, but they cannot be understood as limiting the scope of the invention patent. It should be pointed out that, for ordinary technicians in this field, several variations and improvements can be made without departing from the concept of the present invention, and these all belong to the protection scope of the present invention.

Claims

1. A DNAzyme capable of specifically recognizing and detecting Decapoda iridovirus 1, characterized in that: is a deoxyribonucleotide sequence of DNAzymeMCP-1: The MCP-1 is: 5'-GAAAAGTGTTATCCGGGGAAAGATCATCGTACTGTT ATCTCCGAGCCGGTCGACCAGCATCGGTAGGCGTATCT-3'.

2. The DNAzyme capable of specifically recognizing and detecting Decapoda Iridescent Virus 1 according to claim 1, characterized in that: The DNAzyme sequence has a labeled quencher group at the 3' end.

3. Use of the DNAzyme capable of specifically identifying and detecting decapod iridescent virus 1 as described in any one of claims 1 to 2 in the preparation of a biosensor capable of rapidly detecting decapod iridescent virus 1.

4. A kit capable of specifically identifying and detecting Decapoda iridovirus 1, characterized in that: The kit contains the DNAzyme according to any one of claims 1-2.

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