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Application of molecular marker in diagnosis and treatment of esophageal squamous cell carcinoma

A technology for esophageal squamous cell carcinoma and drugs, applied in the field of biomedicine, can solve problems such as limited application value

Active Publication Date: 2016-08-24
QINGDAO MEDINTELL BIOMEDICAL CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Although markers such as SCC, CEA, CA724, and CA199 have been used clinically, the above molecular markers have limited application value in the clinical diagnosis of esophageal squamous cell carcinoma

Method used

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  • Application of molecular marker in diagnosis and treatment of esophageal squamous cell carcinoma
  • Application of molecular marker in diagnosis and treatment of esophageal squamous cell carcinoma
  • Application of molecular marker in diagnosis and treatment of esophageal squamous cell carcinoma

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0063] Example 1 Screening for gene markers associated with esophageal squamous cell carcinoma

[0064] 1. Sample collection

[0065] Paracancerous tissues and esophageal squamous cell carcinoma tissue samples were collected from 6 cases of esophageal squamous cell carcinoma. All the above specimens were obtained with the consent of the organizational ethics committee.

[0066] 2. RNA sample preparation (using miRNA kit for operation)

[0067] The tissues obtained above were shredded, put into liquid nitrogen and ground into powder, and RNA was extracted and isolated according to the instructions in the kit. details as follows:

[0068] 1) Isolation of RNA:

[0069] A. Add in tissue homogenate or cells Reagent II 1ml;

[0070] B. Stand at room temperature for 3 minutes, add 0.2ml chloroform and shake vigorously for 15 seconds;

[0071] C. Place on ice for 10 minutes;

[0072] D. Centrifuge at 12000g for 15min at 4°C;

[0073] E. Transfer 80% of the aqueous phase in...

Embodiment 2

[0088] Example 2 QPCR sequencing verification of differential expression of MYOC gene

[0089] 1. Large-sample QPCR verification of differential expression of MYOC genes. According to the sample collection method in Example 1, 50 cases of esophageal squamous cell carcinoma adjacent tissues and 50 cases of esophageal squamous cell carcinoma tissues were selected.

[0090] 2. The RNA extraction steps are the same as in Example 1.

[0091] 3. Reverse transcription: use the reverse transcription kit of TAKARA company to operate. Specific steps are as follows:

[0092] (1) Take 1 μg of total RNA for reverse transcription, add 2 μl of Oligo(dT), and mix well. Immediately after 5 min in water bath at 70°C, ice bath for 2 min.

[0093] (2) Construct a 25 μl reaction system, including 5 μl of 5× reverse transcription buffer, 5 μl of dNTP (2.5 mM), 40 U / μl of RNasin, 200 U / μl of M-MLV, and make up to the expected volume with nuclease-free water.

[0094] (3) After 42°C water bath f...

Embodiment 3

[0115] Overexpression of embodiment 3MYOC gene

[0116] 1. Cell culture

[0117] Human esophageal squamous carcinoma cell line KYSE 150 was incubated at 37°C and 5% CO in DMEM containing 10% fetal bovine serum and 1% P / S. 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 and passage.

[0118] 2. Overexpression of MYOC gene

[0119] 2.1 Construction of MYOC gene expression vector

[0120] Amplification primers were designed according to the coding sequence of the MYOC gene (as shown in SEQ ID NO.1), and the primer sequences were as follows:

[0121] Forward primer: 5'-CCGAAGCTTGCCACCATGAGGTTCTTCTGTGCACGT-3' (SEQ ID NO.7)

[0122] Reverse primer: 5'-CGGGCGGCCGCCATCTTGGAGAGCTTGATGTCATA-3' (SEQ ID NO.8)

[0123] The coding sequence of the full-length MYOC gene was amplified from the cDNA library of adult fetal brain (clontech company, catalog number: 638831). The...

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Abstract

The invention discloses application of a molecular marker in diagnosis and treatment of esophageal squamous cell carcinoma. The molecular marker is MYOC gene. Experiments prove that the MYOC gene has differential expression in esophageal squamous cell carcinoma tissues and may function as an index for early diagnosis of esophageal squamous cell carcinoma. It is also proved that it is possible to inhibit the growth and proliferation of esophageal squamous cell carcinoma cells by up-regulating expression level of the MYOC gene, and the molecular marker may function as a potential therapy target for esophageal squamous cell carcinoma and can be used for guiding the development of new pharmaceuticals.

Description

technical field [0001] The invention belongs to the field of biomedicine and relates to the application of a molecular marker in the diagnosis and treatment of esophageal squamous cell carcinoma, specifically the molecular marker is MYOC gene. Background technique [0002] Esophageal cancer is one of the most common malignant tumors in humans. It mainly includes two types, esophageal squamous cell carcinoma and esophageal adenocarcinoma. In China, esophageal squamous cell carcinoma is the main type. At present, the incidence of esophageal squamous cell carcinoma is increasing year by year, with high metastasis rate and poor prognosis. Since patients with esophageal squamous cell carcinoma have no obvious symptoms in the early stage of onset, they are generally in the middle and late stages when they go to the doctor, and the 5-year survival rate is only 18-30%. In my country, especially in areas with a high incidence of esophageal squamous cell carcinoma, esophageal squamou...

Claims

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Application Information

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IPC IPC(8): C12Q1/68G01N33/68G01N33/574A61K45/00A61P35/00
Inventor 常鹏刘中胜杨承刚
Owner QINGDAO MEDINTELL BIOMEDICAL CO LTD
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