In-situ hybridization assay kit for mRNA level of premalignant CDK8 (cyclin-dependent kinase 8) as well as assay method and application

A detection kit and in situ hybridization technology, applied in the determination/inspection of microorganisms, biochemical equipment and methods, etc., can solve problems such as non-decreasing mortality, drug resistance of tumor cells, failure of anti-cancer wars, etc., and achieve operational Convenience, high sensitivity, and strong specificity

Inactive Publication Date: 2012-07-25
NATUREGEN BIOTECH SHANGHAI
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  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0003] In 2005, eight institutions including the US Institutes of Health, the Cancer Institute, and the Centers for Disease Control and Prevention made an annual report, reviewing the anti-cancer war launched in 1972. The report believed that human beings had failed in the anti-cancer war, and the conclusion was that cancer died The rate did not decrease, and it listed several factors that caused the failure of the anti-cancer war: 1. Tumor cell heterogeneity (polymorphism); 2. Tumor cell drug resistance; 3. Incomplete design of anti-cancer drugs (animals) unscientific model design), etc.
[0015] In view of the fact that the current clinical diagnosis of cancer (imaging medicine and biochemical indicators are all diagnosed after tumor formation) is a late diagnosis, the treatment is also a late treatment, leading to a treatment model that does not reduce the mortality rate

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  • In-situ hybridization assay kit for mRNA level of premalignant CDK8 (cyclin-dependent kinase 8) as well as assay method and application
  • In-situ hybridization assay kit for mRNA level of premalignant CDK8 (cyclin-dependent kinase 8) as well as assay method and application
  • In-situ hybridization assay kit for mRNA level of premalignant CDK8 (cyclin-dependent kinase 8) as well as assay method and application

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Experimental program
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Effect test

Embodiment 1

[0053] Prepare the in situ hybridization kit of this embodiment according to conventional methods, the kit includes hybridization probes, markers, and instructions designed with the mRNA of the CDK8 gene as the detection target gene, wherein:

[0054] Digoxigenin was selected as the probe label in this embodiment.

[0055] Kit hybridization solution composition:

[0056] digestive juice 100μL / tube 1 tube / box colorless transparent liquid protective fluid 100μL / tube 1 tube / box colorless transparent liquid Pre-hybridization solution 1300μL / tube 2 tubes / box colorless transparent liquid Right-sense hybridization solution 10μL / tube 1 tube / box colorless transparent liquid antisense hybridization solution 10μL / tube 1 tube / box colorless transparent liquid blocking solution 1000μL / tube 1 tube / box colorless transparent liquid Alkaline phosphatase antibody 1 μL / tube 1 tube / box colorless transparent liquid ...

Embodiment 2

[0064] The implementation process of applying the nucleic acid in situ hybridization detection method to the CDK8 gene expression of each group of blood samples:

[0065] 1).Take two specimens to be tested;

[0066] 2). Add 50 ml of digestive solution (100 μL of digestive solution plus 99.9 ml of 1× buffer Ⅰ, which is the concentration used) in a glass tank, preheat in a water bath at 37°C for 10 minutes, put 16 slides in, and treat at 37°C for 12 minutes , and then washed with 1× buffer I for 5 min;

[0067] 3). Use 0.2% protection solution (protection solution 1ml plus 1× buffer , 99ml is the concentration used), washed for 10 minutes, washed with three-distilled water for 5 minutes (the above process was carried out in a glass tank), took out the slide, and let it dry naturally;

[0068] 4). Put the slides into a humidifying box, add 25 μL / slice of pre-hybridization solution (add to the place where there are cells), cover with a cover glass, cover the humidifying box tig...

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Abstract

The invention discloses an in-situ hybridization assay kit, comprising a hybridization probe and a marker. The invention also discloses a method for assaying mRNA of a cyclin-dependent kinase 8 (CDK8) gene closely related with the pathology evolution of premalignant melanoma through in-situ hybridization using the kit, and the method comprises the following steps: (1) contacting RNA to be assayed in a substrate with the hybridization probe to form a hybridization complex under a condition in which the hybridization probe and a target sequence can form a stable hybridization complex; and (2) assaying the hybridization complex. By using the kit and the assay method of the invention, the expression level of the CDK8 gene can be assayed in the mRNA level, the assay is earlier than medical imaging and traditional clinical biochemical detection indexes, and screening the premalignant mRNA level is really realized. In addition, the assay method of the invention is simple and convenient, is low in cost, and is convenient to be popularized and applied in county and district hospitals.

Description

technical field [0001] The present invention relates to the field of biological detection, more specifically, relates to the relevant detection technology related to the change of mRNA expression (pathological evolution process) in the pathological evolution of melanoma. Background technique [0002] According to the information provided by authoritative organizations at home and abroad, there are 2.6 million new cancer cases in my country every year, nearly 2.1 million deaths, and more than 7 million patients. Globally, there are 8 million new cancer patients every year, nearly 8 million deaths, and more than 8,400 patients. Ten thousand people, the number will double by 2020, this is a set of terrible figures. The cost of cancer diagnosis and treatment is getting higher and higher. According to the annual treatment cost of cancer patients 200,000 yuan (may be higher in poor areas, and 200,000 yuan in developed areas), there are more than 7 million patients, and the annual c...

Claims

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Application Information

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IPC IPC(8): C12Q1/68
Inventor 裘霖张玉丽裘建英张云福
Owner NATUREGEN BIOTECH SHANGHAI
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