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Specific primer, probe and kit for detecting human breast cancer
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A detection kit and technology for human breast cancer, applied in the direction of DNA/RNA fragments, recombinant DNA technology, etc., to achieve the effects of good sensitivity and specificity, saving time and improving efficiency
Active Publication Date: 2014-12-03
黑龙江省医学科学院
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The sensitivity of the three examinations was evaluated, and it was found that ultrasonography was the highest, at about 90%, and mammography was the second, at about 52%. A means of cancer screening that is still controversial
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Embodiment 1
[0043] Embodiment 1 A kind of establishment that is used for the specific primer of human breast cancer detection and probe
[0044] The RT-PCR primers and probes for human breast cancer-specific methylation of the present invention are based on the human whole genome sequence published by NCBI (National Center for Biotechnology Information), using Primer Premier5.0 and Methyl Primer Express v1.0 Designed and synthesized by Yingwei Jieji (Shanghai) Trading Co., Ltd.
[0045] Primer 1 (CBX7Forward-primer): 5-YGGGTGGATGGATAAATGGAA-3
[0046] Primer 2 (CBX7Reverse-primer): 5-CAAAACCRAAATCTCCCTAATCC-3
[0047] Probe (CBX7probe): 5-TGCGTCGCGGAGAA-3
[0048] Among them, primer 1 and primer 2 are degenerate primers, R=A / G, Y=C / T.
Embodiment 2
[0049] Example 2 A Real-time PCR methylation detection kit for diagnosing human breast cancer
[0050] Include the following primer sequences and probe sequences:
[0051] Primer 1 (CBX7Forward-primer): 5-YGGGTGGATGGATAAATGGAA-3
[0052] Primer 2 (CBX7Reverse-primer): 5-CAAAACCRAAATCTCCCTAATCC-3
[0053] Probe (CBX7probe): 5-TGCGTCGCGGAGAA-3
[0054] Among them, primer 1 and primer 2 are degenerate primers, R=A / G, Y=C / T.
[0055] Also included in the kit is 10xPCR buffer, 50mM MgCl 2 , 10 μM dNTPs, 5 μ / μl Taq enzyme.
Embodiment 3
[0056] Example 3 Application of the kit of the present invention in the diagnosis of human breast cancer
[0057] The Taq enzyme used in the present invention was purchased from Yingwei Jieji (Shanghai) Trading Co., Ltd.; the DNA extraction kit was AxyPrepTM Multisource Genomic DNA Miniprep Kit (Axygen Scientific, Inc.CA, USA); the vulcanization kit was EZ DNAMethylation kit (Zymo Research, Orange, CA, USA); other reagents were domestic analytical reagents.
[0058] The biological materials used in the present invention are all from the Department of Breast Surgery, Tumor Hospital Affiliated to Harbin Medical University, and all samples are provided with informed consent and ethics certificates signed by the patients themselves.
[0059] one. Tissue extraction DNA process (DNA extraction kit, AxyPrepTM Multisource Genomic DNA Miniprep Kit (Axygen Scientific, Inc.CA, USA))
[0060] 1. Take 20mg of tissue (20 cases of human breast cancer samples; 40 cases of human benign brea...
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Abstract
The invention discloses a specific primer, a probe and a kit for detecting human breastcancer and belongs to the field of disease diagnosis and control. A pair of Real-time (RT) PCR (PolymeraseChain Reaction) primers for performing specific methylation detection and corresponding probes are designed, aiming at a promoter region of a CBX7 gene in a human breastcancergenome. The primers and the probes are used for performing RT-PCR detection on a vulcanized sample to be detected, and then whether the sample to be detected is a human breastcancer sample can be judged based on the amplification result. Compared with the conventional diagnosis method of breast cancer, the method provided by the invention is high in speed, high in flux, excellent in flexibility and specificity, and can perform early diagnosis for the breast cancer well.
Description
technical field [0001] The invention relates to a specific primer, a probe and a kit for diagnosing diseases, in particular to a specific primer, a probe and a kit for detecting human breast cancer, belonging to the field of disease diagnosis and prevention. Background technique [0002] Breast cancer is the most common malignancy in women and a lifestyle disease. Changes in lifestyle, eating habits, surrounding environment, and a series of changes in marriage and childbearing behaviors such as late marriage and childbirth may all lead to an increase in the incidence of breast cancer. According to the latest statistics, there are about 1.5 million new breast cancer patients in the world every year, and 570,000 people die from breast cancer. Breast cancer has become the leading cause of death for women. Although European and American countries are high-incidence areas of breast cancer, the incidence and mortality of breast cancer in my country are increasing year by year. D...
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