Primer set, kit, library and application for multi-gene combined detection of gynecological tumors
A combined detection technology for gynecological tumors, applied in the field of biochemical detection, can solve the problems of expensive instruments, difficult to reduce the cost of building a database experiment, and unfavorable clinical promotion, etc., to achieve the effect of simple process, easy experiment automation, and low false positive rate
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Embodiment 1
[0062] A kit for multi-gene joint detection of gynecological tumors, which specifically includes the following components: a PCR amplification module and a magnetic bead purification module;
[0063] The PCR amplification module includes the following components:
[0064]
[0065] The magnetic bead purification module includes the following components:
[0066] Magnetic beads 90μL / sample;
[0067] ddH 2 O 100μL / sample.
Embodiment 2
[0069] (1) Genomic DNA extraction
[0070] Collect 2 mL of peripheral blood of the subject (the subject is a person who voluntarily undergoes high-end physical examination) and place it in an EDTA anticoagulation tube, and take 300 μL of EDTA anticoagulant blood from the ultra-clean workbench for genomic DNA extraction. According to the salting-out whole blood DNA purification kit (manufacturer: Guangzhou Meiji Biotechnology Co., Ltd., product model: D3311-02) for genomic DNA extraction, and use Nanodrop to determine the DNA concentration, record the corresponding concentration and 260 / 280 and 260 / The ratio of 230. A total of 1 sample was tested in this test.
[0071] (2) The first round of multiple PCR amplification (the PCR amplification module of the kit prepared in Example 1): Using the genomic DNA of the sample to be tested prepared in step (1) as a template, the reagents prepared in Example 1 are used The box is used to prepare multiple PCR amplification reaction system fo...
Embodiment 3
[0086] Collect 2 mL of peripheral blood from the subject (the subject is a person who voluntarily undergoes a high-end physical examination) to extract genomic DNA and perform the first round of multiplex PCR amplification, the second round of PCR amplification, the second round of PCR product purification, quality inspection, and library Quantitative, diluted library, computer-based sequencing, etc.; apply bioinformatics analysis software NextGENe to convert the sequencing data into FASTQ files, and perform data analysis, compare the DNA sequence obtained by sequencing to the reference target gene sequence, and identify 14 carcinogens Genes (AKT1, APC, BRAF, CTNNB1, EGFR, FBXW7, KRAS, NRAS, PIK3CA, PP2R1A, PTEN, TP53, BRCA1, BRCA2) were extracted from SNVs and Indels, and mutations on target genes were obtained by excluding genetic polymorphism sites For information, see Example 2 for specific methods.
[0087] Genetic test results such as Figure 4 As shown in Table 3, the qua...
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