Novel mutant SNP locus of breast cancer treatment gene RBM12B and applications thereof
A breast cancer, site technology, applied in the field of biomedicine, to achieve the effect of convenient and easy diagnosis
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Embodiment 1
[0052] The collection of embodiment 1 sample and the arrangement of sample data
[0053] From January 2010 to December 2015, the inventor collected a large number of blood samples from patients with new breast cancer in Shenzhen Second People's Hospital. After sorting out the sample data, the inventor selected 25 samples that met the following criteria At the same time, 10 healthy women aged 25-55 were selected as controls for whole-exome microarray testing. The sample selection criteria were as follows:
[0054] 1. Breast cancer cases diagnosed by pathology, among which 3 patients have a family history of cancer and are marked as X1, X2, and X3 respectively;
[0055] 2. Have not received radiotherapy or chemotherapy before blood collection, and have no previous history of cancer;
[0056] 3. Healthy female controls matched with the age of the case
[0057] The demographic data and clinical data of these samples were collected systematically.
Embodiment 2
[0058] Example 2 Extraction and Purification of Peripheral Blood DNA
[0059] Among the above-mentioned 25 eligible breast cancer patients and 10 healthy female controls, the age balance of the two groups was comparable.
[0060] The specific steps are:
[0061] 1. Add hemolysis reagent (i.e., lysate, 40 parts) to the peripheral blood stored in a 2mL cryopreservation tube. The volume of the solution was adjusted to 2000mL, the same below), and it was completely transferred after inverting to mix well.
[0062] 2. Removal of red blood cells: Fill the 5mL centrifuge tube to 4mL with hemolysis reagent, mix by inverting, 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.
[0063] 3. Extract DNA: add 1mL extract solution (each 300mL contains 122.5mL0.2M sodium chloride, 14.4mL0.5M ethylenediaminetetraacetic acid, 15mL10% sod...
Embodiment 3
[0068] The whole exome detection of embodiment 3SNP
[0069] The two groups of people in Example 2 were detected by the whole exome chip to obtain relevant results.
[0070] 1. Library construction
[0071] Beijing Novogene Technology Co., Ltd. uses Agilent's liquid-phase chip capture system to efficiently enrich human DNA from the entire exon region, and then perform high-throughput and high-depth sequencing on the Illumina Hiseq platform. The Agilent SureSelect Human All ExonV5 kit was used for library construction and capture experiments, the reagents and consumables recommended in the instructions were strictly used, and the operation was performed according to the latest optimized experimental procedures.
[0072] The basic process of the experiment: Genomic DNA was randomly broken into fragments with a length of 180-280bp by a Covaris crusher, and after end repair and A-tailing, adapters were connected to both ends of the fragments to prepare a DNA library. After pooli...
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