Kit for quantitatively detecting expression level of specific gene 1 mRNA (messenger Ribonucleic Acid) in human breast cancer

A breast cancer and kit technology, applied in the field of kits for quantitatively detecting the expression level of human breast cancer-specific gene 1 mRNA, can solve the problems of no positive rate detection technology, achieve good accuracy and specificity, and high detection specificity , the effect of broad application prospects

Inactive Publication Date: 2014-03-12
ANHUI PROVINCIAL HOSPITAL
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  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0007] The detection of BCSG1 mRNA in the bone marrow and blood of breast cancer patients is of great significance for judging the occurrence, metastasis and recurrence of breast cancer, evaluating the treatment effect and dynamic observation of the disease, but there is no suitable detection technology with a high positive rate at present

Method used

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  • Kit for quantitatively detecting expression level of specific gene 1 mRNA (messenger Ribonucleic Acid) in human breast cancer
  • Kit for quantitatively detecting expression level of specific gene 1 mRNA (messenger Ribonucleic Acid) in human breast cancer
  • Kit for quantitatively detecting expression level of specific gene 1 mRNA (messenger Ribonucleic Acid) in human breast cancer

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0044] Embodiment 1, kit for quantitatively detecting human breast cancer specific gene 1 mRNA expression level

[0045] The composition of the kit and its preparation method are as follows:

[0046] 1. Red blood cell lysate

[0047] The solvent is water, the solute and its concentration in the red blood cell lysate are respectively 1.52 mol / L of ammonium chloride, 0.1 mol / L of potassium bicarbonate and 0.01 mol / L of disodium edetate. The red blood cell lysate has a pH value of 7.2 at 25°C.

[0048] 2. RNase-free water

[0049] Add diethyl pyrocarbonate (DEPC, purchased from Biobasic Company) to deionized water to a final concentration of 0.05% (volume percentage), place at 22-25°C for 10-12 hours, then autoclave at 121°C for 20 minutes , placed at 22-25°C for later use.

[0050] 3. RNA extraction solution

[0051] For Trizol reagent.

[0052] 4. dNTP mixture

[0053] Contains dATP, dCTP, dGTP, and dTTP, and is stored in the form of its sodium salt-water solution. At 25...

Embodiment 2

[0168] Example 2, detection of breast cancer specific gene 1 mRNA expression

[0169] The kit prepared in Example 1 was used to detect the expression level of breast cancer specific gene 1 mRNA in the specimens of the following experimental group and control group.

[0170] Experimental group: 33 cases of pathologically diagnosed breast cancer patients, of which 12 cases have been clinically diagnosed with metastasis.

[0171] Control group: 15 patients with benign breast disease and 10 healthy people.

[0172] 1. Experiment preparation

[0173] 1. Dilute the erythrocyte lysate with sterilized deionized water at a ratio of 1:9 to form the erythrocyte lysate dilution.

[0174] 2. Prepare 25mL of 75% ethanol solution.

[0175] 2. Sampling

[0176] Take 3 mL of fresh venous blood from the subject in a sterile centrifuge tube, and use EDTA as an anticoagulant (1.44 mg / mL whole blood). The sample should be used immediately after collection. If it cannot be used immediately, it...

Embodiment 3

[0218] Embodiment 3, test kit and application thereof

[0219] 1. Kit composition and preparation method

[0220] Same as described in Example 1, the difference is as follows:

[0221] 1. PCR reaction buffer: composed of tris(hydroxymethyl)aminomethane hydrochloride with a final concentration of 17.9mmol / L, magnesium chloride with a final concentration of 2.55mmol / L and potassium chloride with a final concentration of 90.0mmol / L , the solvent is water; wherein, the final concentration is the final concentration of each substance in the PCR reaction buffer.

[0222] 2. PCR reaction solution: composed of the PCR reaction buffer, Taq enzyme, dATP, dCTP, dGTP, dTTP, dUTP, uracil DNA glycosylase (UNG enzyme) and the primer pair and probe of Example 1, the rest is water;

[0223] The concentration of the PCR reaction buffer in the PCR reaction solution is 0.9 μl / μl;

[0224] The concentration of the Taq enzyme in the PCR reaction solution is 0.08U / μl;

[0225] The concentration...

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Abstract

The invention discloses a kit for quantitatively detecting expression level of a specific gene 1 mRNA (messenger Ribonucleic Acid) in human breast cancer. The kit comprises a specific primer pair formed by a primer shown as a sequence 1 in a sequence table and a primer shown as a sequence 2 in the sequence table, and also comprises a specific probe with a nucleotide sequence shown as a sequence 3 in the sequence table. The kit provided by the invention can be used for accurate qualitative and quantitative analysis of human breast cancer specific gene BCSG1 mRNA expression, and the detection specificity is up to 100%. Compared with the current clinical test method, the proliferation of breast cancer cells can be found earlier, and the sensitivity of whole blood detection is 100 copies / ml. The kit provided by the invention can be used for real-time monitoring of the expression level of BCSG1 mRNA in bone marrow and blood of breast cancer patients, has an important significance on determining of incidence, metastasis and relapse of breast cancer, treatment evaluation and dynamic observation of the disease, and has a broad application prospect.

Description

technical field [0001] The invention relates to a kit for quantitatively detecting the expression level of human breast cancer specific gene 1 mRNA. Background technique [0002] Breast cancer is a common malignant tumor that seriously threatens women's life and health. Due to differences in geographical environment, living habits, etc., the incidence of breast cancer in countries around the world varies greatly. Most countries in North America and Northern Europe are high-incidence areas; Most countries are low-incidence areas. In North America, Europe and other developed countries, the incidence of female breast cancer ranks first in the incidence of female malignant tumors. According to the estimates of the American Cancer Society, there are 120,000 new cases of breast cancer in the United States every year, and the incidence rate is 72.2 / 100,000. Although my country is a low-incidence area of ​​female breast cancer in the world, the incidence of breast cancer has incr...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12Q1/68
Inventor 程民胡世莲魏海明沈国栋陈永艳徐维平徐婷娟王卫东
Owner ANHUI PROVINCIAL HOSPITAL
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