Application of FBXO31 gene and related products in preparing gastric cancer diagnostic reagent
A diagnostic reagent and gene technology, applied in the field of bioengineering, can solve problems such as unclear expression level and function of FBXO31
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Embodiment 1
[0060] RT-PCR, qRT-PCR and Western blot showed that the mRNA and protein levels of FBXO31 were significantly decreased in human gastric cancer tissues;
[0061] 1. Materials and methods
[0062] (1) Patient
[0063] 56 cases of gastric cancer and their surrounding tissues were obtained from patients who underwent gastric tumor resection in Shandong Cancer Hospital during 2012-2013, and the specimens were stored at -80°C. The age, gender, pathological type, degree of differentiation, tumor size (diameter), invasion, local and distant metastasis of the patient at the time of surgery were statistically analyzed. The details are shown in Table 1:
[0064] Table 1. Patient and tumor characteristics and the expression level of FBXO31 protein in normal mucosa and gastric cancer tissues
[0065]
[0066]
[0067] 1. Pathological tumor-node-metastasis.
[0068] 2. Histological differentiation status: LDA, poorly differentiated adenocarcinoma; MDA, moderately differentiated a...
Embodiment 2
[0098] The relationship between the expression of FBXO31 detected by immunohistochemistry and the clinical parameters and postoperative survival of patients with gastric cancer
[0099] 1. Materials and methods
[0100] (1) Tissue chip
[0101] Paraffin-embedded human gastric cancer tissue chips were purchased from Shanghai Xinchao Co., Ltd. One chip contains 180 spots, including 90 specimens of gastric cancer patients. Each patient contains two points, one point represents the cancer tissue, and the other point represents the corresponding paracancerous tissue. Information about the patient's sex, age, tumor size, clinical data, and survival period were all provided by the company.
[0102] (2) Immunohistochemical detection
[0103] ① Dewaxing and hydration: Bake tissue sections at 60°C for 2 hours before dewaxing
[0104] a. Xylene dewaxing 5mim×3 times
[0105] b. Soak in absolute ethanol for 5min×2 times
[0106] c. Soak in 95% ethanol for 5min×2 times
[0107] d. ...
Embodiment 3
[0133] Effect of FBXO31 on the proliferation ability and cell cycle progression of gastric cancer cells detected by colony formation assay and flow cytometry
[0134] 1. Materials and methods
[0135] (1) Experimental materials and cell culture
[0136] Human gastric cancer cell lines BGC-823, HGC-27 were purchased from Shanghai Institute of Biochemistry and Cell Biology, Chinese Academy of Sciences.
[0137] Cell culture at 37°C, 95% air, 5% CO 2 constant temperature incubator.
[0138] The medium was 1640 medium (purchased from Invitrogen, Carlsbad, CA) containing 10% fetal bovine serum (FBS), 100 U / mL penicillin and 2 mmol / L L-glutamine.
[0139] Human miR-20a mimics, miR-17 mimics, blank control mimics, miR-20a inhibitors, miR-17 inhibitors, and blank control inhibitors were purchased from Ribobio (Guangzhou, China).
[0140] Both the pCMV-myc-FBXO31 vector and the F-box sequence deletion mutation vector pCMV-myc-FBXO31ΔF were constructed according to the methods in th...
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