A mutated gene for assessing breast cancer risk and its detection kit
A detection kit and breast cancer technology, applied in the field of biomedicine, can solve the problems of affecting the curative effect and prognosis, insufficient detection of BRCA1/2 gene mutation in breast cancer, influence of metabolic pathways, etc., achieving high accuracy, strong sensitivity, and simple method. Effect
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Embodiment 1
[0041] Example 1 Preparation of a detection kit for assessing the risk of breast cancer
[0042] The production and operation process of the kit of the present invention are based on Sanger sequencing, scanning, detection and typing technology. The kit includes DNA extraction reagents; the kit also includes primers for specifically amplifying ARHGEF18:NM_001130955:exon12:c.C2101T gene mutation, such as the sequence of the forward primer is shown in SEQ ID No.1, and the sequence of the reverse primer is It is shown in SEQ ID No.2; the kit can also include reagents commonly used in PCR reactions, such as Taq enzyme, dNTP mixture, MgCl 2 solution, deionized water, etc.; these commonly used reagents are well known to those skilled in the art, and can also include peripheral blood DNA from normal healthy females in the control group. The PCR reaction system used was 25 μL: Taq Buffer 2.5 μL, DNA 1 μL, forward primer 0.5 μL, reverse primer 0.5 μL, 10 mM dNTP 0.5 μL, Taq enzyme 0.2 ...
Embodiment 2
[0054] Example 2 Use of the detection kit prepared in Example 1
[0055] 1. Extraction of DNA
[0056] Blood samples from 46 patients with clinically detected breast cancer were taken and DNA was extracted.
[0057] The specific steps are:
[0058] (1) Add hemolysis reagent (i.e. lysate, 40 parts) to the peripheral blood stored in the 2mL cryopreservation tube. Dilute the volume of TrisHcl solution to 2000mL, the same below), mix by inverting and transfer completely.
[0059] (2) Removal of red blood cells: Fill the 5mL centrifuge tube to 4mL with hemolysis reagent, mix by inversion, centrifuge at 4000rpm for 10 minutes, and discard the supernatant. Add 4 mL of hemolysis reagent to the precipitate, invert and wash again, centrifuge at 4000 rpm for 10 minutes, and discard the supernatant.
[0060] (3) DNA extraction: Add 1 mL of extract solution (each 300 mL contains 122.5 mL of 0.2 M sodium chloride, 14.4 mL of 0.5 M ethylenediamine tetraacetic acid, 15 mL of 10% sodium do...
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