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Primer set, reagent and kit for detecting methylation of RPRM gene and PRDM5 gene and use thereof

A technology of methylation and primer set, applied in the biological field, can solve the problems such as inability to obtain pathological diagnosis, and achieve the effect of easy mastery, strong specificity and high sensitivity

Active Publication Date: 2018-06-01
JIANGSU MICRODIAG BIOMEDICINE TECH CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The detection of serum PG level is a good indicator to reflect the function and status of gastric mucosa, and has the advantages of non-invasive, simple and fast; gastroscopy detection is considered to be the "gold standard" for gastric cancer detection, and its sensitivity and specificity are both over 90%. Intuitive and most effective means of early detection of gastric cancer; before the use of endoscopy, barium meal radiography was the main means of diagnosis and screening of gastric cancer, but it was gradually replaced by endoscopic detection because it could not obtain pathological diagnosis

Method used

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  • Primer set, reagent and kit for detecting methylation of RPRM gene and PRDM5 gene and use thereof
  • Primer set, reagent and kit for detecting methylation of RPRM gene and PRDM5 gene and use thereof
  • Primer set, reagent and kit for detecting methylation of RPRM gene and PRDM5 gene and use thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0088] The composition of embodiment 1 kit and the processing of sample to be tested

[0089] 1. The kit for simultaneously detecting methylation of RPRM gene and PRDM5 gene provided by the present invention includes a DNA processing kit and a methylation gene detection kit. Among them, the composition of the DNA processing kit is shown in Table 1, and the composition of the methylated gene detection kit is shown in Table 2.

[0090] Table 1 DNA processing kit

[0091]

[0092]

[0093] Table 2 Methylation gene detection kit

[0094] Reagent

main ingredient

PCR Mix

dNTPs, primers, buffer salts

Taq Enzyme Mix

Taq enzyme, probe, buffer salt

negative control

TE buffer, unmethylated human genomic DNA

positive control substance

TE buffer, methylated human genomic DNA

[0095] 2. Instruments and equipment: The instruments and equipment required for testing are as follows:

[0096]Vortex shaker, rotary mixer, 15mL ...

Embodiment 2

[0123] Example 2 Detection of RPRM and PRDM5 methylated genes

[0124] 1) According to the reaction system in Table 3, prepare the PCR pre-reaction solution, mix it and distribute it into PCR tubes, add the sample to be tested, the reference substance and purified water as templates, and the volume of each reaction is 20 μL;

[0125] Table 3 PCR reaction system

[0126]

[0127] 2) Perform fluorescent quantitative PCR reaction according to the reaction conditions in Table 4.

[0128] Table 4 reaction conditions

[0129]

[0130] 3) After the sample is put on the machine, save the experiment name, run the PCR reaction, and write the sample layout and sample name according to the experiment layout in time.

Embodiment 3

[0131] Embodiment 3PCR result analysis

[0132] 1. in Select "Analysis" in the 480 basic software module column.

[0133] 2. Select the "Abs Quant / Fit Points" analysis mode.

[0134] 3. Select "Filter Comb 465-510".

[0135] 4. Select the "Cycle Range" window, set "First Cycle" to "1"; "Last Cycle" to "50".

[0136] 5. In the "Cycle Range" window, set the background to "5-22", click the blue "Background" button, set the "Min Offset" to "4", and set the "Max Offset" to "twenty one".

[0137] 6. Select the "Noise Band" window, set "Noise Band" to "Noise Band (Fluoresc)", and manually change the value of "Noise Band" to "2.0".

[0138] Note: If the background value is higher than 2.0, the baseline can be adjusted according to the actual situation, that is, the value of "Noise Band", until the amplification curve without obvious exponential growth period should be negative.

[0139] 7. Select the "Analysis" window, "Threshold(Auto)" will automatically adjust the "Threshold"...

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Abstract

The invention provides a primer set, reagent and kit for detecting methylation of a RPRM gene and a PRDM5 gene and a use thereof and relates to the technical field of biotechnology. Each one of the primer set, reagent and kit for simultaneously detecting methylation of the RPRM gene and the PRDM5 gene comprises a primer pair for detecting a RPRM gene and a primer pair for detecting a PRDM5 gene, has high specificity and high sensitivity and realizes simple, fast and accurate simultaneous detection of the methylation states of the RPRM and PRDM5 genes. The invention provides a method for simultaneously detecting methylation of the RPRM gene and the PRDM5 gene. The method is easy to operate and learn. The invention also provides a use of the primer set, reagent or kit in a product for assistant diagnosis and / or early screening of gastric cancer.

Description

technical field [0001] The invention relates to the field of biotechnology, in particular to a primer set, reagent, kit and application for detecting the methylation of RPRM gene and PRDM5 gene. Background technique [0002] Gastric cancer is one of the most common tumors in the world. Its incidence rate ranks fourth among all malignant tumors, and its death rate ranks second among malignant tumors. More than 70% of gastric cancer cases exist in the developing stage. nation. There are about 400,000 new gastric cancer cases in my country every year, accounting for 40% of the world's total incidence. In my country, the early diagnosis rate of gastric cancer is less than 10%. Once diagnosed, most of them are in the middle and advanced stages, and the 5-year survival rate is less than 20%. Up to 60% or more. Therefore, how to realize early detection of gastric cancer is the key to reduce its mortality. [0003] More than 90% of gastric cancer is adenocarcinoma. In 1965, Laure...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/6886C12N15/11
CPCC12Q1/6886C12Q2600/154
Inventor 王弢张田田王冬华
Owner JIANGSU MICRODIAG BIOMEDICINE TECH CO LTD
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