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Detection kit for diagnosis of patients with stomach cancer based on multiple genes

A kit and genetic technology, applied in the biological field, to achieve the effect of high sensitivity

Pending Publication Date: 2017-05-03
BEIJING EXELLON MEDICAL TECH CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

As a result, the frequency of false negative tests is very high, leading patients and health care workers to believe that the disease does not exist when, in fact, the patient may have a serious cancer that requires immediate attention
Moreover, use of these markers may give false positive signals in up to 1 / 3 of individuals with benign gastric disease

Method used

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  • Detection kit for diagnosis of patients with stomach cancer based on multiple genes
  • Detection kit for diagnosis of patients with stomach cancer based on multiple genes
  • Detection kit for diagnosis of patients with stomach cancer based on multiple genes

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0031] Embodiment one: DNA extraction

[0032] DNA extraction reagent consists of lysis buffer, washing buffer and elution buffer, see Table 1; lysis buffer consists of protein denaturant, detergent, pH buffer and nuclease inhibitor; protein denaturant: hydrochloric acid Guanidine (purchased from sigma company); detergent is: Tween20 (purchased from sigma company); pH buffer is: Tris-HCl (purchased from Aladdin company); nuclease inhibitor is: EDTA (purchased from sigma company) .

[0033] Table 1 Reagents used for DNA extraction

[0034]

[0035] In this embodiment, a plasma sample (purchased from Beijing 301 Hospital) is taken as an example to extract plasma circulating tumor DNA. The extraction method includes the following steps: (1) Take 1ml of plasma, add the same volume of lysis buffer, and then add proteinase K and CarrierRNA, Make the final concentrations of 100mg / L and 1μg / ml respectively, shake and mix, and incubate at 55°C for 30min; (2) Add 100μl magnetic beads...

Embodiment 2

[0036] Example 2: Bisulfite treatment of DNA

[0037] Bisulfite treatment of DNA is to use bisulfite reagent to process, bisulfite reagent is composed of bisulfite buffer and protection buffer; bisulfite buffer is sodium bisulfite (purchased from Sigma ) and water mixed liquid; protection buffer is an oxygen radical scavenger hydroquinone (purchased from sigma company) and water mixed liquid. The protein denaturant was guanidine hydrochloride (purchased from sigma); the pH buffer was Tris-HCl (purchased from Aladdin); the nuclease inhibitor was EDTA (purchased from sigma). The reagent formula of this embodiment is shown in Table 2.

[0038] Table 2 Reagents used for bisulfite treatment of DNA

[0039]

[0040] In Example 2, the DNA extracted in Example 1 was used as the treatment object, and bisulfite was used to treat the DNA. The specific methods included: (1) preparing a bisulfite buffer solution, weighing 1 g of sodium bisulfite powder, and adding water to prepare in...

Embodiment 3

[0041] Example 3: Detection of DNA methylation by fluorescent quantitative PCR

[0042] In this embodiment, the detection genes are p16, CDH1, MGMT, RARB and RNF180 genes, and the internal reference gene is ACTB. The internal reference primer pair of the internal reference gene ACTB is a primer pair composed of DNA shown in sequence 46 of the sequence listing and the DNA shown in sequence 47 of the sequence listing; the nucleotide sequence of the internal reference probe of the internal reference gene ACTB is shown in sequence 48 of the sequence listing . In this example, the DNA treated with bisulfite in Example 2 is used as a template for PCR amplification.

[0043] The final concentration of the fluorescent quantitative PCR amplification reaction system is composed of: 1 × PCR buffer (purchased from NEB Company), 0.5 mM dNTPs (purchased from NEB Company), 0.5 μM target gene detection primer, 0.2 μM target gene fluorescent probe, 1 μM Target gene blocking primer, 0.3 μM in...

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Abstract

The invention relates to a detection kit for diagnosis of patients with stomach cancer based on multiple genes. The kit comprises a primer probe composition a, a primer probe composition b, a primer probe composition c, a primer probe composition d and a primer probe composition e; the primer probe composition a comprises a specific primer pair a, a closed primer a and a probe a; the primer probe composition b comprises a specific primer pair b, a closed primer b and a probe b; the primer probe composition c comprises a specific primer pair c, a closed primer c and a probe c; the primer probe composition d comprises a specific primer pair d, a closed primer d and a probe d; the primer probe composition e comprises a specific primer pair e, a closed primer e and a probe e. The kit provided by the invention can be applied to diagnosis of stomach cancer, provides a new fast, reliable and accurate method for diagnosis of stomach cancer, and plays an important role in the medicinal detection field.

Description

technical field [0001] The invention relates to the field of biotechnology, in particular to a detection kit for diagnosing gastric cancer patients based on multiple genes. Background technique [0002] In my country, the incidence of gastrointestinal system tumors ranks first, among which gastric cancer is the highest. The annual incidence of gastric cancer in the world is about 870,000, and there are more than 300,000 in my country. In 1962, the Japanese Gastric Cancer Research Association formally proposed the concept of early gastric cancer (EGC), which was a milestone event in the diagnosis and treatment of gastric cancer. Due to the concept of early gastric cancer, it became possible to achieve clinical cure and long-term survival of gastric cancer. After half a century of research, diagnosis and treatment, the understanding of early gastric cancer has gradually deepened, and the diagnosis of early gastric cancer has gradually become more refined. Especially in recent...

Claims

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Application Information

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IPC IPC(8): C12Q1/68
CPCC12Q1/6886C12Q2600/112C12Q2600/118C12Q2600/166
Inventor 李明明索伟克蒲珏
Owner BEIJING EXELLON MEDICAL TECH CO LTD
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