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Breast cancer susceptibility genes BRCA1 and BRCA2 whole-gene trap primers, kit and method

A susceptibility gene and whole gene technology, applied in the field of breast cancer susceptibility genes BRCA1 and BRCA2 whole gene capture primers, can solve problems such as affecting gene function and affecting RNA shearing, and achieve low cost, simple operation, and uniform sequencing data. good effect

Pending Publication Date: 2018-03-06
HANGZHOU D A GENETIC ENG
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  • Summary
  • Abstract
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  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] At present, the detection of BRCA1 and BRCA2 focuses on the exons, but some studies have also shown that the variation of the nucleic acid sequence on the introns will also affect the splicing of RNA, thereby affecting the function of the gene

Method used

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  • Breast cancer susceptibility genes BRCA1 and BRCA2 whole-gene trap primers, kit and method

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Experimental program
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Effect test

Embodiment 1

[0025] Step 1: DNA Extraction

[0026] Take 200 μL of peripheral blood, and perform DNA extraction according to the instructions of the whole blood genomic DNA extraction kit (purchased from Hangzhou Xinjie Biotechnology Co., Ltd.), and extract DNA for subsequent experiments or store at -20°C.

[0027] Step 2: PCR Amplification and Detection

[0028] (1) Amplify BRCA1 and BRCA2 primer pairs, the nucleotide sequences of which are shown in SEQ ID NO.: 1-32;

[0029] (2) In the PCR 8 connected tubes, prepare the following reaction system:

[0030] Total volume 20 μL, containing 4 μL of 5× PrimeSTAR GXL Buffer, 1.6 μL of dNTP Mixture (2.5 mM each), 1 μL of PrimeSTAR GXL DNA Polymerase, upstream primers (SEQ ID NO.: 1, 3, 5, 7, 9, 11, 13, 15, 17, 19, 21, 23, 25, 27, 29) 0.4 μM, downstream primers (SEQ ID NO.: 2, 4, 6, 8, 10, 12, 14, 16, 18, 20, 22, 24 ,26,28,30) 0.4μM, template DNA 50-100ng, deionized water to make up the volume to 20μL. The amplification program in the PCR rea...

Embodiment 2

[0034] Step 1: DNA Extraction

[0035] Take 200 μL of peripheral blood, and perform DNA extraction according to the instructions of the whole blood genomic DNA extraction kit (purchased from Hangzhou Xinjie Biotechnology Co., Ltd.), and extract DNA for subsequent experiments or store at -20°C.

[0036] Step 2: PCR Amplification and Detection

[0037] (1) Amplify BRCA1 and BRCA2 primer pairs, the nucleotide sequences of which are shown in SEQ ID NO.: 1-32;

[0038] (2) In the PCR 8 connected tubes, prepare the following reaction system:

[0039] Total volume 20 μL, containing 4 μL of 5× PrimeSTAR GXL Buffer, 1.6 μL of dNTP Mixture (2.5 mM each), 1 μL of PrimeSTAR GXL DNA Polymerase, upstream primers (SEQ ID NO.: 1, 3, 5, 7, 9, 11, 13, 15, 17, 19, 21, 23, 25, 27, 29) 0.4 μM, downstream primers (SEQ ID NO.: 2, 4, 6, 8, 10, 12, 14, 16, 18, 20, 22, 24 ,26,28,30) 0.4μM, template DNA 50-100ng, deionized water to make up the volume to 20μL. The amplification program in the PCR rea...

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Abstract

The invention provides nucleotide sequences for amplification testing of HER-2 gene copy number, a kit and application thereof. The nucleotide sequences contain 8 pairs of PCR amplification primers for amplification of BRCA1 (Gene ID: 672) full-length gene and 8 pairs PCR amplification primers for amplification of BRCA2 (Gene ID: 675) full-length gene. DNA trapped by amplification can be used forsubsequent high-throughput sequencing analysis. In comparison with a probe targeting trapping technology and a multiplex PCR technology, the method of the invention is simple to operate, is low-cost,has good sequencing data uniformity, can reach more than 99% coverage with small sequencing data amount, and is helpful for providing gene detection technical support for risk assessment or medicationof genetic tumors related with breast cancer susceptibility genes and establishment of a database.

Description

technical field [0001] The invention belongs to the technical field of gene detection, and in particular relates to a complete gene capture primer, a kit and applications of breast cancer susceptibility genes BRCA1 and BRCA2. Background technique [0002] Both BRCA1 and BRCA2 belong to tumor suppressor genes, which are two genes related to breast and ovarian cancer susceptibility, and are mainly involved in DNA damage repair. BRCA1 and BRCA2 are involved in homologous recombination, DNA damage repair, embryonic growth, and transcriptional regulation. BRCA1 and BRCA2 gene mutations are the cause of most autosomal dominant familial hereditary breast cancer and ovarian cancer, especially BRCA2 gene mutation is more closely related to male breast cancer. BRCA gene mutation carriers have a significantly higher lifetime risk of breast cancer and are prone to early onset. Breast cancer is the most common malignant tumor in women. In 2007, there were 1.3 million new breast cancer ...

Claims

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Application Information

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IPC IPC(8): C12Q1/686C12N15/11
CPCC12Q1/686C12Q2537/143
Inventor 任绪义张锋周韵王裕舟赵铃铃
Owner HANGZHOU D A GENETIC ENG
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