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Application of reagent for detecting expression level of FEN1 gene to preparation of reagents used for diagnosing and predicting invasion and metastasis of liver cancer

A technology of gene expression levels and reagents, applied in the biological field, to achieve the effect of promoting transfer

Inactive Publication Date: 2018-11-16
THE SECOND AFFILIATED HOSPITAL OF CHONGQING MEDICAL UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

So far, there are no good biomarkers that can be used for early diagnosis and prediction of tumor invasion and metastasis

Method used

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  • Application of reagent for detecting expression level of FEN1 gene to preparation of reagents used for diagnosing and predicting invasion and metastasis of liver cancer
  • Application of reagent for detecting expression level of FEN1 gene to preparation of reagents used for diagnosing and predicting invasion and metastasis of liver cancer
  • Application of reagent for detecting expression level of FEN1 gene to preparation of reagents used for diagnosing and predicting invasion and metastasis of liver cancer

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0064] Example 1 qPCR detection of the expression of FEN1 gene in liver cancer cells and normal liver cells

[0065] 1 sample collection:

[0066] Culture normal liver cell HL7702 and a series of liver cancer cells SK-HEP-1, MHCC97-H, HepG2, SMMC-7721, HCCLM3, Huh7. DMEM medium containing 10% fetal bovine serum and 1% double antibody in 37%, 5% CO 2 , Cultivated in an incubator with a relative humidity of 90%. Change the medium once every 2-3 days, and use 0.25% EDTA-containing trypsin for routine digestion.

[0067] 2 RNA sample preparation:

[0068] Cellular RNA was extracted using QIAGEN's cellular RNA extraction kit, and the operation was performed according to the specific steps in the manual.

[0069] 3 reverse transcription:

[0070] Use a 25μL reaction system, take 1ug total RNA from each sample as template RNA, and add the following components to the PCR tube: DEPC water, 5X reverse transcription buffer, 10mM dNTP, 0.1mM DTT, 30uM Oligo dT, 200U / μL M-MLV, templa...

Embodiment 2

[0080] Example 2 Using qPCR to detect the expression of FEN1 in patients with liver cancer and adjacent tumors

[0081] 1 The cancer tissues and corresponding paracancerous tissues of 16 pairs of liver cancer patients were collected, and the patients gave informed consent, and all the above specimens were obtained with the consent of the ethics committee.

[0082] 2 RNA sample preparation:

[0083] Tissue RNA was extracted using QIAGEN tissue RNA extraction kit, and the operation was performed according to the specific steps in the manual.

[0084] 3 reverse transcription:

[0085] Concrete steps are with embodiment 1.

[0086] 4 results:

[0087] The result is as figure 2 As shown in Fig. A, where the expression of FEN1 in 16 pairs of liver cancer patients and adjacent tumors was detected by qPCR, and Fig. B is the statistical result. Compared with adjacent tissues, the expression of FEN1 gene was up-regulated in liver cancer tissues, and the difference was statisticall...

Embodiment 3

[0088] Example 3 Using immunohistochemistry to detect the expression of FEN1 in the cancer and adjacent tumors of liver cancer patients

[0089] 1 Tissue block fixed, dehydrated, embedded

[0090] 16 pairs of liver cancer and paracancerous tissues were fixed in 4% paraformaldehyde for 3-4 hours, and tissues of appropriate size were put into embedding boxes for dehydration. Enter in order: 75% alcohol 1.5h, 95% alcohol 1.5h, 95% alcohol 1h, absolute ethanol 1.5h, absolute ethanol 1h, xylene No. 1 0.5h, xylene No. 2 0.5h. Turn on the embedding machine, use an iron lunch box to hold the paraffin block and put it into the tissue tank of the embedding machine to heat and dissolve. Put the dehydrated tissue together with the embedding box into the tissue tank of the machine in turn, and then put it into the paraffin lunch box No. 1 for the second wax dipping, and then paraffin lunch box No. 2 for three times of wax dipping. Then drop liquid paraffin into the mold, take out the soa...

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Abstract

The invention especially relates to the application of a reagent for detecting the expression level of the FEN1 gene to preparation of reagents used for diagnosing and predicting the invasion and metastasis of liver cancer and the application of a FEN1 gene inhibitor to preparation of drugs used for inhibiting the invasion and metastasis of liver cancer, belonging to the field of biotechnology. Itis found for the first time that the FEN1 gene is related to the development and progression of liver cancer; interference experiment results in the invention prove that the change of the expressionlevel of the FEN1 gene can influence the invasion and metastasis of cells; and animal experiment results show that the expression of the FEN1 gene can promote the metastasis of liver cancer.

Description

technical field [0001] The invention relates to the field of biotechnology, in particular to the application of a reagent for detecting the expression level of fen1 gene in the preparation of a reagent for diagnosing and predicting invasion and metastasis of liver cancer. Background technique [0002] Primary liver cancer ranks third among the most lethal tumors in the world in terms of lethality. However, early detection of liver cancer is difficult, and the high recurrence and metastasis rates after liver cancer resection have seriously affected the treatment effect and survival time of liver cancer patients. The etiology and pathogenesis of primary liver cancer have not been determined. At present, it is believed that it is related to chemical carcinogens such as liver cirrhosis, viral hepatitis, and aflatoxin and environmental factors. Clinical manifestations mainly include the following aspects: [0003] 1. Pain in the liver area: more than half of the patients had pa...

Claims

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Application Information

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IPC IPC(8): C12Q1/6886G01N33/574A61K45/00A61P35/04
CPCA61K45/00A61P35/04C12Q1/6886C12Q2600/158G01N33/57438G01N33/57484
Inventor 汤慧李传飞胡鹏任红
Owner THE SECOND AFFILIATED HOSPITAL OF CHONGQING MEDICAL UNIV
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