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Breast cancer combined diagnosis markers and detecting kit

A breast cancer and kit technology, applied in the fields of biochemistry and molecular biology, can solve problems such as inapplicable clinical disease diagnosis, and achieve the effects of avoiding excessive tissue biopsy, high specificity and sensitivity, and prolonging survival

Active Publication Date: 2016-01-20
王义明
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, most of the current molecular diagnosis studies focus on the relationship between gene polymorphisms and breast cancer, which can only give the susceptibility risk of patients, and are not suitable for the diagnosis of clinical diseases

Method used

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  • Breast cancer combined diagnosis markers and detecting kit
  • Breast cancer combined diagnosis markers and detecting kit
  • Breast cancer combined diagnosis markers and detecting kit

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0056] Embodiment 1 breast cancer detection and / or curative effect evaluation kit

[0057] Kit 1

[0058] Including 8 systems, namely RECQL, RECQL4, RECQL5 and GAPDH, the real-time fluorescence quantitative PCR reaction system and the positive template system of these 4 genes, among which,

[0059] 19.0 μL of real-time fluorescent quantitative PCR reaction system for each gene, including 10 μL of PCR reaction buffer, 0.8 μL of each gene amplification primer with a concentration of 10 μM, 1.0 μL of a gene-specific probe with a concentration of 2 μM, and 7.2 μL of RNase-freewater;

[0060] The positive template system for each gene included a concentration of 1 x 10 9 copies / μL of the purified gene amplification product.

[0061] RECQL upstream primer: 5'-ACAAAGGGCAATCAGGAATCA-3';

[0062] RECQL downstream primer: 5'-CATTGGCTGACCATTTTCTATGAAC-3';

[0063] RECQL probe sequence: 5'-AATTCATGCAGGTGCTTACCATGCCAA-3';

[0064] RECQL4 upstream primer: 5'-TCTCTCCCCTGCTGTCACTCA-3'; ...

Embodiment 2

[0085] Embodiment 2 The using method of kit of the present invention

[0086] (1) Real-time fluorescent quantitative PCR reaction

[0087] The kit of the present invention can also only include primers and probes of four genes, and positive templates of amplification products of four genes, and then assist the existing real-time fluorescent quantitative PCR kit to perform detection together.

[0088] Single gene real-time fluorescent quantitative PCR reaction system

[0089] Reagent

Volume (μL)

PCR reaction buffer

10

Primer F / R (10μM)

0.8

Probe (2μM)

1.0

cDNA

1.0

RNase-free water

7.2

total capacity

20

[0090] Note: The above reaction reagents use Taqman Fluorescence Quantitative Kit from ABI Company in the United States, and the same type of products from other companies can also be used.

[0091] The PCR amplification conditions were: pre-denaturation at 95°C for 10 min, denaturation at ...

Embodiment 3

[0099] The establishment of three target gene standard curves and method repeatability of embodiment 3:

[0100] (1) Preparation of standard curve

[0101] Template: use the quantitative positive template of the kit of the present invention to draw a quantitative standard curve to detect the repeatability of the kit.

[0102] Instrument: Model 7300 real-time fluorescent quantitative PCR instrument of American ABI Company.

[0103] Method: The positive templates of three target genes (RECQL, RECQL4, RECQL5) and one housekeeping gene (GAPDH) were diluted 10-fold, and then diluted to 10 7 、10 6 、10 5 、10 4 and 10 3 copies / μL. Take 1 μL each for real-time fluorescent quantitative PCR reaction.

[0104] See attached for standard curve figure 1 , the RECQL standard curve equation is: y=-3.8833x+41.323, the correlation coefficient R 2 =0.9969; RECQL4 standard curve equation is: y=-4.036x+37.778, correlation coefficient R 2 =0.9990; RECQL5 standard curve equation is: y=-3.95...

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Abstract

The invention discloses a breast cancer combined diagnosis markers and a detecting kit. Studies show that RECQL, RECQL4 and RECQL5 genes can serve as the combined markers and used for preparing breast cancer detection and / or therapeutic evaluation reagents. According to the kit formed by the markers, a specificity RT-PCR primer designed with GAPDH serving as a reference gene, and a probe, a real-time fluorescence quantitative PCR method is used for measuring the content of mRNA of the RECQL, RECQL4 and RECQL5 genes drifting away in peripheral blood monouclear cells and urine of a patient, and diagnosis and therapeutic effect evaluation can be carried out on breast cancer. Compared with a conventional diagnosis method, the method has the advantages that trauma is small, use is fast, throughput is high, sensitivity and specificity are good, and early diagnosis and therapeutic evaluation can be carried out on the breast cancer better.

Description

technical field [0001] The invention relates to the fields of biochemistry and molecular biology, in particular to a combined diagnostic marker and detection kit for breast cancer. Background technique [0002] The burden of cancer in China is increasing, with 1.6 million people diagnosed with cancer and 1.2 million people dying of cancer every year. Like many other countries, breast cancer is the most common cancer among women in China. Globally, China accounts for 12.2% of newly diagnosed breast cancer cases and 9.6% of breast cancer deaths. [0003] Breast cancer is a systemic disease, the incidence rate is increasing year by year, and the age of onset is gradually getting younger. As a highly heterogeneous tumor, lymphatic or hematogenous metastasis can occur in the early stage, and it is one of the common malignant tumors in women. [0004] Although the incidence of breast cancer in China is currently low, since the 1990s, the incidence rate of breast cancer in China h...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/68C12N15/11
CPCC12Q1/6886C12Q2600/158C12Q2600/178C12N15/11C12Q1/68
Inventor 王义明罗国安
Owner 王义明
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