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Application of CYLD, IFFO1 and downstream target gene in comprehensive treatment and detection of drug resistance of ovarian cancer

A comprehensive detection and comprehensive treatment technology, applied in the direction of recombinant DNA technology, microbial measurement/testing, DNA/RNA fragments, etc., can solve problems such as rare research reports, and achieve the effect of promoting expression increase and drug resistance increase

Inactive Publication Date: 2022-06-28
ZHENGZHOU UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, in tumor drug resistance, there are few research reports

Method used

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  • Application of CYLD, IFFO1 and downstream target gene in comprehensive treatment and detection of drug resistance of ovarian cancer
  • Application of CYLD, IFFO1 and downstream target gene in comprehensive treatment and detection of drug resistance of ovarian cancer
  • Application of CYLD, IFFO1 and downstream target gene in comprehensive treatment and detection of drug resistance of ovarian cancer

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0034] We collected tissues from 18 ovarian cancer patients and 6 normal ovarian tissues, and detected the expression level of CYLD in the specimens of each patient by qRT-PCR. By comparing the expression level of CYLD in patients with normal ovarian tissue, it was found that the expression level of CYLD in the specimens of ovarian cancer patients was significantly lower, and was far lower than the expression level of CYLD in normal ovarian tissue. Therefore, we believe that CYLD plays an inhibitory role in the occurrence and development of ovarian cancer. At the same time, we detected the expression level of CYLD protein in patient tissue by western bloting in patient tissue samples, and obtained the same results.

Embodiment 2

[0036]In ovarian cancer platinum drug-sensitive cell line OVCAR3 and platinum drug-resistant cell line OVCAR3-DDP, western blotting was used to detect the protein expression level of CYLD; specific methods:

[0037] 1. Extraction of total cell protein

[0038] 2. SDS-PAGE electrophoresis

[0039] 3. Semi-dry transfer film

[0040] 4. Immunoblotting

[0041] After transferring the membrane, check whether the marker has been transferred to the membrane (or Ponceau red staining) to determine the transfer effect. After washing the membrane with TBST, 5% nonfat dry milk (prepared in TBST) was blocked overnight at 4°C. Add primary anti-CYLD polyclonal antibody (1:1000) (purchased from CST Company), shake at room temperature for 2h. Rinse 3 times with TBST, 10 min each time. Add secondary antibody (1:200) (purchased from cell signal company), shake at room temperature for 1 h. Rinse 3 times with TBST, 10 min each time.

[0042] 5. ECL detection

[0043] 6. Analysis of results...

Embodiment 3

[0045] The OVCAR ovarian cancer platinum drug-sensitive cell line that stably inhibits CYLD expression and the OVCAR-DDP ovarian cancer platinum drug-resistant cell line that stably overexpress CYLD were constructed by lentiviral packaging and puromycin screening. Then, the stable expression in stable cell lines OVCAR / shNC, OVCAR / shCYLD1, OVCAR / shCYLD3, and ovarian cancer platinum drug-sensitive cell line OVCAR3, platinum drug-resistant cell line OVCAR3-DDP and drug-resistant cell line were detected by western bloting. The expression level of CYLD in OVCAR3-DDP CYLD of CYLD was verified after three times to ensure that the silencing of CYLD and the construction of overexpression cell lines were completed. Through western blotting, it was found that the expression of CYLD decreased significantly in the cell line that silenced CYLD; in another cell line OVCAR-DDP CYLD, it was found that the protein level of CYLD increased significantly after overexpression of CYLD. After the ver...

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Abstract

The invention belongs to the technical field of tumor drug resistance detection application, and relates to application of a detection kit for simultaneously detecting CYLD and IFFO1 as well as downstream target genes Bax, Bcl-XL, Bc13 and ABCB1 to detect generation of tumor drug resistance. The invention particularly relates to application of CYLD and IFFO1 in discovering platinum drug resistance of ovarian cancer.

Description

technical field [0001] The invention belongs to the technical field of tumor drug resistance detection and application, and in particular relates to a method for simultaneous detection of CYLD and IFFO1 and downstream target genes ABCB1, Bcl-XL, Bax and Bcl3 for comprehensive treatment of ovarian cancer drug resistance and a comprehensive detection kit. Develop applications. Background technique [0002] Ovarian cancer is a disease that seriously threatens women's health. Its incidence is only the third in gynecological malignant tumors, but its mortality rate ranks first. According to the latest data, its incidence is increasing year by year. trend, while the mortality rate did not improve significantly. The current treatment principle for ovarian cancer is as follows: on the basis of surgical resection (R0 resection) without residual as much as possible, it is supplemented by platinum-based chemotherapy. However, due to the location of the organs in the abdominal cavity,...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/6886C12N15/11
CPCC12Q1/6886C12Q2600/106C12Q2600/158
Inventor 张烨王琳汪伟江秉华贾小玉
Owner ZHENGZHOU UNIV