A biomarker for predicting liver cancer MVI and a detection kit

Abstract

The present application belongs to the field of biomarker detection, and particularly relates to a biomarker for predicting liver cancer MVI and a detection kit. The present application detects biomarkers AFP, GP73, OPN and CST1 which can be used for predicting liver cancer MVI. The principle of the detection kit obtained by the present application is that the to-be-detected biomarker has high affinity with the corresponding apt, when the to-be-detected biomarker exists, the cDNA is released, and as the content of the to-be-detected biomarker increases, the release amount of the cDNA increases, after magnetic separation, the cDNA is left in the solution, at this time, the H1-H2 double-stranded complex is added, the cDNA can induce the opening of the ring structure of the H1 chain and the H2 chain, the H1 chain and the H2 chain are tightly combined, at this time, the cDNA is released again, after multiple cycles, the amplification of the fluorescence signal is realized, and thus the existence of trace biomarkers can be detected, and the accuracy is high.

Description

Technical Field

[0001] This invention belongs to the field of biomarker detection, specifically relating to a biomarker and detection kit for predicting MVI in liver cancer. Background Technology

[0002] Liver cancer is a relatively common cancer, and while there are many effective treatments available, such as surgery, liver transplantation, and radiofrequency ablation, the recurrence rate remains high. Microvascular invasion (MVI) is a hallmark of tumor invasiveness, indicating a high risk of recurrence after liver resection. It is defined as a tumor lesion located within a blood vessel, as observed under a microscope. Small branches of the portal vein are more susceptible to tumor invasion, followed by the hepatic veins, and sometimes it can also occur in branches of the hepatic artery, bile ducts, and lymphatic vessels. Using a "7-point" baseline sampling method, pathologically, tumors can be classified into M0 (no MVI), M1 (≤5 MVI lesions located in the liver tissue near the lesion), and M2 (>5 MVI lesions, or located in the liver tissue far from the lesion). The "7-point" baseline sampling method has been validated for its scientific validity. MVI is one of the most important risk factors for intrahepatic recurrence in patients undergoing radical liver cancer surgery, especially M2 grade MVI, which is a high-risk factor for postoperative recurrence and intrahepatic metastasis. For liver cancer patients at high risk of MVI, alternative treatment strategies and preoperative adjuvant therapy should be considered. Therefore, it is necessary to establish non-invasive and accurate models to predict MVI grading, thereby assisting surgeons in selecting appropriate surgical methods for patients.

[0003] Common methods for predicting MVI include magnetic resonance imaging (MRI), a non-invasive medical imaging technique widely used in the diagnosis and treatment of diseases in various fields, including liver cancer. However, most of these studies use MVI negative / positive classification, and there are relatively few studies on specific MVI grading predictions, thus providing relatively limited information. Summary of the Invention

[0004] Based on the above problems, this invention provides a biomarker for predicting MVI in liver cancer, specifically alpha-fetoprotein, Golgi protein 73, osteopontin, and serum cysteine ​​protease inhibitor 1. A detection kit is also prepared for detection. The biomarker and its corresponding apt have high affinity. When the biomarker is present, cDNA is released, and the amount of cDNA released increases with the increase of the biomarker concentration. After magnetic separation, the cDNA remains in solution. When the H1-H2 double-stranded complex is added, the cDNA induces the opening of the circular structure of the H1 and H2 chains, and the H1 and H2 chains bind tightly. The cDNA is then released again. Through multiple cycles, the fluorescence signal is amplified. Experiments demonstrate that the detection kit obtained by this invention has high detection accuracy.

[0005] This invention provides a biomarker for predicting MVI in liver cancer, comprising the following substances: Alpha-fetoprotein (AFP), Golgi protein 73 (GP73), osteopontin (OPN), and serum cysteine ​​protease inhibitor 1 (CST1).

[0006] The present invention also provides a detection kit for predicting MVI in liver cancer, comprising the following components by weight: 4-8 parts of MNPs-apt-cDNA probe and 3-6 parts of H1-H2 double-stranded complex. The MNPs-apt-cDNA probe is obtained by linking streptavidin-modified MNPs with biotin-modified apts. The MNPs are magnetic nanoparticles, preferably iron oxide nanoparticles, and the apts are aptamers for biomarkers. Preferably, the apt and cDNA sequences involved in the MNPs-apt-cDNA probe are as follows: AFP-apt: 5'-TCAGGTGCAGTTCTCGACTCGGTCTTGATGTGGGTCCCCCC-3'; GP73-apt: 5'-GCAGTTGATCCTTTGGATACCCTGG-3'; OPN-apt: 5'-TGTGTGCGGCACTCCAGTCTGTTACGCCGC-3'; CST1-apt:5'-ACGTAGAGGTGGGCCGCACCATAT-3'; AFP-cDNA: 5'-CACATCAAGACCG-3'; GP73-cDNA: 5'-ATCAAAGGA-3'; OPN-cDNA: 5'-AACAGACTGGAGTG-3'; CST1-cDNA: 5'-GCGGCCCACCT-3'.

[0007] The sequences corresponding to the above apt are shown as SEQ ID NO:1-SEQ ID NO:8, respectively.

[0008] The preparation method of the MNPs-apt-cDNA probe is as follows: L1. At a volume ratio of 1:1, streptavidin-modified MNPs and biotin-modified apt were obtained. The mixture was shaken at room temperature for 30-50 min and washed 2-3 times with buffer 1 to obtain the MNPs-apt complex. L2. Add the MNPs-apt complex obtained in step L1 to the cDNA solution and shake at room temperature for 20-40 min. After the treatment, magnetically separate to obtain the MNPs-apt-cDNA complex. Dissolve the MNPs-apt-cDNA complex in buffer 2. The mass-volume ratio of MNPs-apt-cDNA complex to buffer 2 is 1-1.5 mg: 0.8-1 mL. After mixing, the MNPs-apt-cDNA probe is obtained.

[0009] Preferably, in step L1, the concentration of streptavidin-modified MNPs is preferably 1 μM, the concentration of biotin-modified apt is preferably 1 mg / μL, and the buffer 1 is prepared as follows: 100 mM sodium chloride, 5 mM magnesium chloride, and 20 mM Tris-HCl.

[0010] Preferably, in step L2, the mass-to-volume ratio of MNPs-apt complex to cDNA solution is 2-5 mg: 1-3 mL, the concentration of cDNA solution is preferably 1 μM, and buffer 2 is prepared as follows: 2.5 mM magnesium chloride, 150 mM sodium chloride, 10 mM potassium chloride, and 10 mM Tris-HCl.

[0011] The preparation method of the H1-H2 double-chain complex is as follows: S1. Dissolve the H1 chain in Tris-HCl buffer solution to obtain a 4 μM H1 chain solution. Weigh the H1 chain solution, silver nitrate solution, ammonium acetate solution, and sodium borohydride solution according to a volume ratio of 2-3:1:8-12:1-1.2. Shake the H1 chain solution, silver nitrate solution, and ammonium acetate solution to mix well. Let it stand for 1-2 hours in the dark. After that, add sodium borohydride solution and stir at 400-500 rpm for 20-30 minutes. After that, let it stand for 24-36 hours in the dark to obtain a cyclized H1 chain solution. S2. Prepare a cyclized H2 chain solution following the steps in S1; S3. Mix the cyclized H1 chain solution obtained in step S1 with the cyclized H2 chain solution obtained in step S2 at a volume ratio of 1:1 to obtain the H1-H2 double-chain complex.

[0012] Preferably, in step S1, the pH of the Tris-HCl buffer solution is 7.4, the concentration is preferably 10 mM, the concentration of the silver nitrate solution is 200 μM, the concentration of the ammonium acetate solution is 240 μM, and the concentration of the sodium borohydride solution is 100 μM.

[0013] Preferably, the sequences of the H1 and H2 chains corresponding to the biomarker are as follows: AFP-H1 chain: 5'-CCCCCCCCCATACGGTCTTGATGTGGGGTGGGGTGCCTACTAGAGT-3'; AFP-H2 chain: 5'-TAGTAGGCACCCCACCCACACATCGGGTGGGGTGGGGTGGG-3'; GP73-H1 chain: 5'-CCCCCCCCCATATCCTTTGGATGGGTGGGGTGGCTTCTAGGTC-3'; GP73-H2 chain: 5'-AGAAGCCACCCCACCCATCCAAGGGTGGGGGTGGGGTGGG-3'; OPN-H1 chain: 5'-CCCCCCCCCATACACTCCAGTCTGTTGGGTGGGGTGACTTGCCGG-3'; OPN-H1 chain: 5'-AAGTCACCCCACCCAACAGACGGGTGGGGTGGGGTGGG-3'; CST1-H1 chain: 5'-CCCCCCCCCATAAGGTGGGCCGCGGGTGGGGTGCGTAGGCGG-3'; CST1-H2 chain: 5'-CTACGCACCCCACCCGCGGCGGGTGGGTGGGGTGGG-3'.

[0014] The beneficial effects of this invention are as follows: This invention detects alpha-fetoprotein (AFP), Golgi protein 73 (GLP-73), osteopontin, and serum cysteine ​​protease inhibitor 1 (CPI1), biomarkers that can be used to predict microvascular vitiligo (MVI) in liver cancer. The principle of the detection kit obtained by this invention is as follows: the biomarker to be tested has a high affinity for its corresponding aptamer. When the biomarker is present, cDNA is released, and the amount of cDNA released increases with the concentration of the biomarker. After magnetic separation, the cDNA remains in solution. When the H1-H2 double-stranded complex is added, the cDNA induces the opening of the circular structure of the H1 and H2 chains, and the H1 and H2 chains bind tightly. The cDNA is then released again. Through multiple cycles, the fluorescence signal is amplified, thereby detecting the presence of trace amounts of the biomarker. Attached Figure Description

[0015] The accompanying drawings, which are included to provide a further understanding of this application and form part of this application, illustrate exemplary embodiments and are used to explain this application, but do not constitute an undue limitation of this application. In the drawings: Figure 1 This is a graph showing the detection results of the biomarker AFP; Figure 2 The image shows the detection results for the biomarker GP73. Figure 3 This is a graph showing the detection results of the biomarker OPN. Figure 4 This is a graph showing the detection results of the biomarker CST1. Detailed Implementation

[0016] To more clearly illustrate the overall concept of this application, a detailed description is provided below with reference to the accompanying drawings and embodiments. Numerous specific details are set forth in the following description to provide a more thorough understanding of the invention. However, it will be apparent to those skilled in the art that the invention can be practiced without one or more of these details. In other instances, certain technical features well-known in the art have not been described to avoid confusion with the invention.

[0017] Example 1: This example provides a detection kit for predicting MVI in liver cancer, the details of which are as follows: The detection kit for predicting MVI in liver cancer in this embodiment contains the following components by weight: 4 parts of MNPs-apt-cDNA probe and 3 parts of H1-H2 double-stranded complex. The MNPs-apt-cDNA probe is obtained by linking streptavidin-modified MNPs with biotin-modified apts. MNPs are iron oxide nanoparticles, and apts are aptamers for biomarkers. Preferably, the apt and cDNA sequences involved in the MNPs-apt-cDNA probe are as follows: AFP-apt: 5'-TCAGGTGCAGTTCTCGACTCGGTCTTGATGTGGGTCCCCCC-3'; GP73-apt: 5'-GCAGTTGATCCTTTGGATACCCTGG-3'; OPN-apt: 5'-TGTGTGCGGCACTCCAGTCTGTTACGCCGC-3'; CST1-apt: 5'-ACGTAGAGGTGGGCCGCACCATAT-3'; AFP-cDNA: 5'-CACATCAAGACCG-3'; GP73-cDNA: 5'-ATCAAAGGA-3'; OPN-cDNA: 5'-AACAGACTGGAGTG-3'; CST1-cDNA: 5'-GCGGCCCACCT-3'; The preparation method of the MNPs-apt-cDNA probe is as follows: L1. At a volume ratio of 1:1, streptavidin-modified MNPs (1 μM) and biotin-modified apt (1 mg / μL) were obtained. The mixture was shaken at room temperature for 30 min and washed twice with buffer 1. Buffer 1 was prepared as follows: 100 mM sodium chloride, 5 mM magnesium chloride, and 20 mM Tris-HCl to obtain the MNPs-apt complex. L2. Add the MNPs-apt complex obtained in step L1 to the cDNA solution (1 μM). The mass-to-volume ratio of MNPs-apt complex to cDNA solution is 2 mg: 1 mL. Shake at room temperature for 20 min. After the treatment, magnetically separate to obtain the MNPs-apt-cDNA complex. Dissolve the MNPs-apt-cDNA complex in buffer 2. The buffer 2 is prepared as follows: 2.5 mM magnesium chloride, 150 mM sodium chloride, 10 mM potassium chloride, 10 mM Tris-HCl. The mass-to-volume ratio of MNPs-apt-cDNA complex to buffer 2 is 1 mg: 0.8 mL. After mixing, the MNPs-apt-cDNA probe is obtained. The preparation method of the H1-H2 double-chain complex is as follows: S1. Dissolve the H1 chain in Tris-HCl buffer solution (pH 7.4, 10 mM) to obtain a 4 μM H1 chain solution. Weigh the H1 chain solution, silver nitrate solution (200 μM), ammonium acetate solution (240 μM), and sodium borohydride solution (100 μM) according to a volume ratio of 2:1:8:1. Shake the H1 chain solution, silver nitrate solution, and ammonium acetate solution to mix well. Let it stand for 1 h in the dark. After that, add sodium borohydride solution and stir at 400 rpm for 20 min. After that, let it stand for 24 h in the dark to obtain a cyclized H1 chain solution. S2. Prepare a cyclized H2 chain solution following the steps in S1; S3. Mix the cyclized H1 chain solution obtained in step S1 with the cyclized H2 chain solution obtained in step S2 at a volume ratio of 1:1 to obtain the H1-H2 double-chain complex. The sequences of the H1 and H2 chains corresponding to the biomarkers are as follows: AFP-H1 chain: 5'-CCCCCCCCCATACGGTCTTGATGTGGGGTGGGGTGCCTACTAGAGT-3'; AFP-H2 chain: 5'-TAGTAGGCACCCCACCCACACATCGGGTGGGGTGGGGTGGG-3'; GP73-H1 chain: 5'-CCCCCCCCCATATCCTTTGGATGGGTGGGGTGGCTTCTAGGTC-3'; GP73-H2 chain: 5'-AGAAGCCACCCCACCCATCCAAGGGTGGGGGTGGGGTGGG-3'; OPN-H1 chain: 5'-CCCCCCCCCATACACTCCAGTCTGTTGGGTGGGGTGACTTGCCGG-3'; OPN-H1 chain: 5'-AAGTCACCCCACCCAACAGACGGGTGGGGTGGGGTGGG-3'; CST1-H1 chain: 5'-CCCCCCCCCATAAGGTGGGCCGCGGGTGGGGTGCGTAGGCGG-3'; CST1-H2 chain: 5'-CTACGCACCCCACCCGCGGCGGGTGGGGGTGGGGTGGG-3'.

[0018] Example 2: This example provides a detection kit for predicting MVI in liver cancer, the details of which are as follows: The detection kit for predicting MVI in liver cancer in this embodiment contains the following components by weight: 5 parts of MNPs-apt-cDNA probe and 4 parts of H1-H2 double-stranded complex. The preparation method of the MNPs-apt-cDNA probe is as follows: L1. At a volume ratio of 1:1, streptavidin-modified MNPs (1 μM) and biotin-modified apt (1 mg / μL) were obtained. The mixture was shaken at room temperature for 40 min, washed 3 times with buffer 1 (the same as in Example 1) to obtain the MNPs-apt complex. L2. Add the MNPs-apt complex obtained in step L1 to the cDNA solution (1 μM). The mass-to-volume ratio of MNPs-apt complex to cDNA solution is 3 mg: 2 mL. Shake at room temperature for 30 min. After the treatment, magnetically separate to obtain the MNPs-apt-cDNA complex. Dissolve the MNPs-apt-cDNA complex in buffer 2. The buffer 2 is the same as in Example 1. The mass-to-volume ratio of MNPs-apt-cDNA complex to buffer 2 is 1.2 mg: 0.9 mL. After mixing, the MNPs-apt-cDNA probe is obtained. The preparation method of the H1-H2 double-chain complex is as follows: S1. Dissolve the H1 chain in Tris-HCl buffer solution (pH 7.4, 10 mM) to obtain a 4 μM H1 chain solution. Weigh the H1 chain solution, silver nitrate solution (200 μM), ammonium acetate solution (240 μM), and sodium borohydride solution (100 μM) in a volume ratio of 2.5:1:10:1.1. Shake the H1 chain solution, silver nitrate solution, and ammonium acetate solution to mix well. Let it stand for 1.5 h in the dark. After that, add sodium borohydride solution and stir at 450 rpm for 25 min. After that, let it stand for 28 h in the dark to obtain a cyclized H1 chain solution. S2. Prepare a cyclized H2 chain solution following the steps in S1; S3. Mix the cyclized H1 chain solution obtained in step S1 with the cyclized H2 chain solution obtained in step S2 at a volume ratio of 1:1 to obtain the H1-H2 double-chain complex. The sequences of atp, cDNA, H1 chain, and H2 chain involved in this embodiment are the same as in Embodiment 1.

[0019] Example 3: The detection kit for predicting MVI in liver cancer in this example contains the following components by weight: 8 parts of MNPs-apt-cDNA probe and 6 parts of H1-H2 double-stranded complex. The preparation method of the MNPs-apt-cDNA probe is as follows: L1. At a volume ratio of 1:1, streptavidin-modified MNPs (1 μM) and biotin-modified apt (1 mg / μL) were obtained. The mixture was shaken at room temperature for 50 min, washed 3 times with buffer 1 (the same as in Example 1) to obtain the MNPs-apt complex. L2. Add the MNPs-apt complex obtained in step L1 to the cDNA solution (1 μM). The mass-to-volume ratio of MNPs-apt complex to cDNA solution is 5 mg: 3 mL. Shake at room temperature for 40 min. After the treatment, magnetically separate to obtain the MNPs-apt-cDNA complex. Dissolve the MNPs-apt-cDNA complex in buffer 2. The buffer 2 is the same as in Example 1. The mass-to-volume ratio of MNPs-apt-cDNA complex to buffer 2 is 1.5 mg: 1 mL. After mixing, the MNPs-apt-cDNA probe is obtained. The preparation method of the H1-H2 double-chain complex is as follows: S1. Dissolve the H1 chain in Tris-HCl buffer solution (pH 7.4, 10 mM) to obtain a 4 μM H1 chain solution. Weigh the H1 chain solution, silver nitrate solution (200 μM), ammonium acetate solution (240 μM), and sodium borohydride solution (100 μM) in a volume ratio of 3:1:12:1.2. Shake the H1 chain solution, silver nitrate solution, and ammonium acetate solution to mix well. Let it stand for 2 h in the dark. After that, add sodium borohydride solution and stir at 500 rpm for 30 min. After that, let it stand for 36 h in the dark to obtain a cyclized H1 chain solution. S2. Prepare a cyclized H2 chain solution following the steps in S1; S3. Mix the cyclized H1 chain solution obtained in step S1 with the cyclized H2 chain solution obtained in step S2 at a volume ratio of 1:1 to obtain the H1-H2 double-chain complex. The sequences of atp, cDNA, H1 chain, and H2 chain involved in this embodiment are the same as in Embodiment 1.

[0020] The sequences involved in Examples 1-3 were all synthesized by Sangon Biotech (Shanghai) Co., Ltd.

[0021] Example 4: This example provides a method for using a detection kit for predicting MVI in liver cancer, as detailed below: Serum samples were homogenized in extraction buffer (volume ratio 1:3) at room temperature for 15 min. The extraction buffer was prepared as follows: 2 mM sodium bicarbonate, 50 mM sodium chloride, and 10 mM EDTA. The samples were then incubated at 4 °C for 6 h and centrifuged at 8000 g for 20 min. The supernatant was collected, and the supernatant and MNPs-apt-cDNA probe were weighed at a volume ratio of 5:1. The samples were incubated at 37 °C for 15 min. After incubation, the samples were magnetically separated, and the separated liquid was collected. The liquid and H1-H2 double-stranded complex were weighed at a volume ratio of 1:0.1. The samples were incubated at 37 °C for 10 min, and the fluorescence spectrum from 615 nm to 750 nm was collected. The excitation wavelength was 495 nm.

[0022] Experimental testing: Kit detection accuracy: Obtain pure AFP, GP73, OPN, and CST1, set up gradient concentrations, and perform tests using the kits obtained in Examples 1-3, respectively, following the method in Example 4: The results are as follows Figures 1-4 As shown, by Figure 1 , Figure 2 , Figure 3 , Figure 4 It can be seen that the kits obtained in Examples 1-3 all have good detection effects on the biomarkers AFP, GP73, OPN and CST1 used to predict MVI of liver cancer. The results of the linear fitting curves are shown in Table 1: Table 1. Linear fitting curves for biomarker detection

[0023] The above description is merely a few embodiments of the present invention and is not intended to limit the present invention in any way. Although the present invention has been disclosed above with reference to preferred embodiments, it is not intended to limit the present invention. Any modifications or alterations made by those skilled in the art without departing from the scope of the technical solution of the present invention using the disclosed technical content are equivalent to equivalent implementation cases and fall within the protection scope of the technical solution of the present invention.

Claims

1. A detection kit for predicting MVI in liver cancer, characterized in that, The test kit is used to detect biomarkers alpha-fetoprotein, Golgi protein 73, osteopontin, and serum cysteine ​​protease inhibitor 1. The kit contains the following components by weight: 4-8 parts of MNPs-apt-cDNA probe and 3-6 parts of H1-H2 double-stranded complex. The MNPs-apt-cDNA probe is obtained by linking streptavidin-modified MNPs with biotin-modified apts. MNPs are magnetic nanoparticles, and apts are aptamers for biomarkers. The apt and cDNA sequences involved in the MNPs-apt-cDNA probe are as follows: AFP-apt: 5'-TCAGGTGCAGTTCTCGACTCGGTCTTGATGTGGGTCCCCCC-3'; GP73-apt: 5'-GCAGTTGATCCTTTGGATACCCTGG-3'; OPN-apt: 5'-TGTGTGCGGCACTCCAGTCTGTTACGCCGC-3'; CST1-apt: 5'-ACGTAGAGGTGGGCCGCACCATAT-3'; AFP-cDNA: 5'-CACATCAAGACCG-3'; GP73-cDNA: 5'-ATCAAAGGA-3'; OPN-cDNA: 5'-AACAGACTGGAGTG-3'; CST1-cDNA: 5'-GCGGCCCACCT-3'; The preparation method of the MNPs-apt-cDNA probe is as follows: L1. At a volume ratio of 1:1, streptavidin-modified MNPs and biotin-modified apt were obtained, shaken at room temperature for 30-50 min, washed with buffer 1, and the MNPs-apt complex was obtained. L2. Add the MNPs-apt complex obtained in step L1 to the cDNA solution and shake at room temperature. After the treatment, magnetic separation is performed to obtain the MNPs-apt-cDNA complex. Dissolve the MNPs-apt-cDNA complex in buffer 2. The mass-volume ratio of MNPs-apt-cDNA complex to buffer 2 is 1-1.5 mg: 0.8-1 mL. After mixing, the MNPs-apt-cDNA probe is obtained. The preparation method of the H1-H2 double-chain complex is as follows: S1. Dissolve the H1 chain in Tris-HCl buffer solution to obtain a 4 μM H1 chain solution. Weigh the H1 chain solution, silver nitrate solution, ammonium acetate solution, and sodium borohydride solution according to a volume ratio of 2-3:1:8-12:1-1.

2. Shake the H1 chain solution, silver nitrate solution, and ammonium acetate solution to mix well. Let it stand for 1-2 hours in the dark. After that, add sodium borohydride solution and stir. Let it stand for 24-36 hours in the dark to obtain a cyclized H1 chain solution. S2. Prepare a cyclized H2 chain solution following the steps in S1; S3. Mix the cyclized H1 chain solution obtained in step S1 with the cyclized H2 chain solution obtained in step S2 at a volume ratio of 1:1 to obtain the H1-H2 double-chain complex. The sequences of the H1 and H2 chains corresponding to the biomarkers are as follows: AFP-H1 chain: 5'-CCCCCCCCCATACGGTCTTGATGTGGGGTGGGGTGCCTACTAGAGT-3'; AFP-H2 chain: 5'-TAGTAGGCACCCCACCCACACATCGGGTGGGGTGGGGTGGG-3'; GP73-H1 chain: 5'-CCCCCCCCCATATCCTTTGGATGGGTGGGGTGGCTTCTAGGTC-3'; GP73-H2 chain: 5'-AGAAGCCACCCCACCCATCCAAGGGTGGGGGTGGGGTGGG-3'; OPN-H1 chain: 5'-CCCCCCCCCATACACTCCAGTCTGTTGGGTGGGGTGACTTGCCGG-3'; OPN-H2 chain: 5'-AAGTCACCCCACCCAACAGACGGGTGGGGTGGGGTGGG-3'; CST1-H1 chain: 5'-CCCCCCCCCATAAGGTGGGCCGCGGGTGGGGTGCGTAGGCGG-3'; CST1-H2 chain: 5'-CTACGCACCCCACCCGCGGCGGGTGGGTGGGGTGGG-3'.

2. The detection kit for predicting MVI in liver cancer according to claim 1, characterized in that, In step L1, the concentration of streptavidin-modified MNPs was 1 μM, and the concentration of biotin-modified apt was 1 mg / μL.

3. The detection kit for predicting MVI in liver cancer according to claim 2, characterized in that, In step L2, the mass-to-volume ratio of MNPs-apt complex to cDNA solution is 2-5 mg: 1-3 mL, and the concentration of cDNA solution is 1 μM.

4. The detection kit for predicting MVI in liver cancer according to claim 3, characterized in that, In step S1, the Tris-HCl buffer solution has a pH of 7.4 and a concentration of 10 mM, the silver nitrate solution has a concentration of 200 μM, the ammonium acetate solution has a concentration of 240 μM, and the sodium borohydride solution has a concentration of 100 μM.

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