Liver cancer diagnosis and treatment marker, namely LINC02128

A liver cancer and liver cancer treatment technology, applied in the field of biomedicine, can solve problems such as functions that need to be further clarified, and achieve the effect of improving accuracy

Inactive Publication Date: 2020-08-11
TAIZHOU MUNICIPAL HOSPITAL
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Compared with protein coding sequences and microRNA, lncRNA research is still in its infancy, and its function needs to be further elucidated

Method used

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  • Liver cancer diagnosis and treatment marker, namely LINC02128
  • Liver cancer diagnosis and treatment marker, namely LINC02128
  • Liver cancer diagnosis and treatment marker, namely LINC02128

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0062] Example 1 QPCR detection of differential expression of LINC02128

[0063] 1. RNA extraction

[0064] Tissue RNA was extracted using QIAGEN tissue RNA extraction kit, and the specific steps in the instructions were followed.

[0065] 2. Reverse transcription reaction

[0066] Use 25μl reaction system, take 1μg total RNA for each sample as template RNA, and add the following components to PCR tubes: DEPC water, 5× reverse transcription buffer, 10mM dNTP, 0.1mM DTT, 30μM Oligo dT, 200U / μl M-MLV, template RNA. Incubate at 42°C for 1 hour, then centrifuge briefly at 72°C for 10 minutes.

[0067] 3. Real-time fluorescent quantitative PCR

[0068]PCR amplification was performed using cDNA as a template. The upstream primer of LINC02128 is 5′-CAGGAGCCAGAGAATCAT-3′ (SEQ ID NO.2), the downstream primer is 5′-TCACGGAGGTTTACCAAAT-3′ (SEQ ID NO.3); GAPDH is an internal reference, and the upstream primer is 5′-CCGGGAAACTGTGGCGTGATGG-3′ (SEQ ID NO.4), the downstream primer is 5'...

Embodiment 2

[0073] Example 2 Effect of LINC02128 gene on the proliferation of liver cancer cells

[0074] 1. Cell culture and cell transfection

[0075] Construct the LINC02128 overexpression vector according to conventional techniques, and the general steps are as follows: amplify LINC02128 to obtain the cDNA sequence, insert it into the eukaryotic expression vector pcDNA3.1(-), construct the LINC02128-pcDNA3.1(-) recombinant vector, and insert It transforms competent cells, picks clones, and double-enzyme cuts identification to obtain LINC02128-pcDNA3.1(-) expression vector containing complete and correct LINC02128 sequence. Plasmids were prepared in large quantities using an endotoxin-free kit.

[0076] Human hepatoma cell SMMC-7721 cells were inoculated in DMEM medium containing 10% fetal bovine serum in 5% CO 2 , Cultured in a constant temperature incubator at 37°C. 24h before transfection press 3×10 5 Inoculate each well in a 6-well plate, and add 2 mL of serum-free medium to ma...

Embodiment 3

[0083] Example 3 Soft agar colony formation experiment

[0084] 1. Experimental steps

[0085] (1) Digest the cells with 0.25% trypsin, pipette gently to make a single cell suspension, and collect the cell pellet by centrifugation.

[0086] (2) Resuspend with DMEM complete medium containing 20% ​​fetal bovine serum, count after appropriate dilution, and adjust the cell concentration to 5×10 3 pieces / ml.

[0087] (3) Prepare two low-melting-point agarose solutions with concentrations of 1.2% and 0.7%, respectively, and maintain them in a 40° C. water bath after autoclaving.

[0088] (4) Mix 1.2% agarose and 2×DMEM medium 1:1, add 2×antibiotics and 20% calf serum, take 3ml of the mixed solution and inject it into a 6cm-diameter plate, let it cool and solidify for 5 minutes, and place it as the bottom agar CO 2 Reserve in the incubator.

[0089] (5) Mix 0.7% agarose and 2×DMEM medium 1:1 in a sterile test tube, then add 0.2ml to the tube with a concentration of 5×10 3 cells...

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Abstract

The invention discloses a liver cancer diagnosis and treatment marker, namely, LINC02128. The invention discloses the LINC02128 which can be used as a biomarker for liver cancer diagnosis and can alsobe used as a new target for developing a medicine for treating liver cancer.

Description

technical field [0001] The invention belongs to the field of biomedicine and relates to LINC02128, a marker for diagnosis and treatment of liver cancer. Background technique [0002] Primary liver cancer is one of the most common malignant tumors in the world, its incidence ranks sixth among all cancers, and its mortality rate ranks second among all cancers. According to pathological classification, primary liver cancer can be divided into different types such as hepatocellular carcinoma (HCC), intrahepatic cholangiocarcinoma, and mixed hepatocellular carcinoma-intrahepatic cholangiocarcinoma. More than 90% of primary liver cancers are hepatocellular carcinomas. The occurrence of liver cancer is a multi-step and complicated process, mostly based on the chronic inflammation of the liver and the changed liver microenvironment, the mutation of many oncogenes or tumor suppressor genes, especially the gradual changes in their expression, and eventually induce liver cancer. Bec...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/6886C12N15/11A61K45/00A61K31/7105A61P35/00A61P1/16
CPCA61K31/7105A61K45/00A61P1/16A61P35/00C12Q1/6886C12Q2600/136C12Q2600/158C12Q2600/178
Inventor 王冬国杨林军陈佳玉
Owner TAIZHOU MUNICIPAL HOSPITAL
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