Liver cancer relative gene DLK1 and its use

A liver cancer, sequence technology, applied in the fields of molecular biology and genetic engineering, can solve the problems of little knowledge and poor prognosis of liver cancer

Inactive Publication Date: 2007-03-14
CHINESE NAT HUMAN GENOME CENT AT SHANGHAI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

Liver cancer is known as the "national disease" in my country. After half a century of exploration, although there is a certain understanding of the early diagnosis and treatment

Method used

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  • Liver cancer relative gene DLK1 and its use
  • Liver cancer relative gene DLK1 and its use
  • Liver cancer relative gene DLK1 and its use

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0036] Extraction of liver tissue RNA

[0037] The RNA extraction reagent uses TRIzol reagent (GIBCO / BRL), which is produced based on the one-step extraction method with acidic phenol. The utensils and water used for RNA extraction must be RNase-free to ensure an RNase-free environment in the experiment.

[0038] Dry-bake the pestle, homogenizer and other utensils at 200°C for 4 hours to remove RNase and cool down; add liquid nitrogen to pre-cool, take the tissue out of the liquid nitrogen quickly, and grind it into powder; use a spatula to put the tissue into the pre- Add TRIzol reagent to the homogenizer and homogenize for several minutes; transfer the homogenized liquid into an RNase-free centrifuge tube, add chloroform, and centrifuge at 4°C to separate layers; transfer the upper aqueous phase to an RNase-free In a centrifuge tube, add isopropanol and centrifuge at 4°C to precipitate RNA; wash the precipitate twice with 75% ethanol; dissolve the precipitate with RNase-fre...

Embodiment 2

[0041] cDNA synthesis

[0042] reverse transcription (RT)

[0043] Total RNA 1 μg

[0044] oligo dT (10pmol / l) 1μl

[0045] h 2 O make up to 12 μl

[0046] Denature at 70°C for 3 minutes, then place on ice to cool;

[0047] Then add 5×Buffer 4μl

[0048] DTT 2μl

[0049] dNTP 1μl

[0050] SuperScript II 1μl

[0051] React at 42°C for 1 hour, denature at 70°C for 15 minutes, and cool on ice.

Embodiment 3

[0053] PCR amplification

[0054]Using β-actin as internal control, the components in the reaction mixture are: β-actin(F) primer, β-actin(R) primer, DLK1(F) primer, DLK1(R) primer, 10×Buffer, MgCl 2 , dNTP, Taq DNA polymerase, and cDNA template are 0.2, 0.2, 0.4, 0.4, 1.0, 1.0, 0.2, 0.1 and 5 μL cDNA template, and finally add ddH 2 O to make the reaction volume 10 μL. The reaction conditions of PCR were denaturation at 94°C for 5min; then each cycle was 30s at 94°C, 30s at 53°C, and 30s at 72°C for a total of 30 cycles; and finally extended at 72°C for 7min.

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Abstract

The present invention discloses human gene Delta like 1 homolog (DLK1) and the application of its coded protein in diagnosing and treating liver cancer. The human DLK1 gene located in 14q32 of chromosome encodes one protein containing 383 amino acids. The human DLK1 gene has expression in liver cancer more obviously than in nearby tissues. Researching on the liver cancer relative human LLK1 gene and its coded protein in depth may find out new mechanism of liver cancer. At the same time, they may become the new target site for liver cancer treatment and may provide important reference for developing liver cancer treating medicine.

Description

technical field [0001] The present invention relates to the fields of molecular biology and genetic engineering, in particular, the present invention relates to the application of a human liver cancer-related gene DLK1 (Delta like 1 homolog) and its coding sequence. Background technique [0002] At present, there are about 12 million patients with chronic hepatitis in my country, and about 300,000 people die of liver disease every year, 50% of which are primary liver cancer, accounting for about 45% of the death toll of liver cancer in the world. The vast majority are related to HBV and HCV infection. The incidence rate of liver cancer ranks 2-3 in my country, mainly in young and middle-aged men, and the incidence rate in East China is significantly higher than that in other regions. In recent years, it has continued to rise. In Shanghai, the incidence of liver cancer ranks third, second only to lung cancer and gastric cancer. The latest data show that this ratio is still ...

Claims

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Application Information

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IPC IPC(8): A61K38/16A61K48/00A61P35/00C12Q1/68
Inventor 黄健张新韩泽广
Owner CHINESE NAT HUMAN GENOME CENT AT SHANGHAI
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