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Human plasma RASSF1A gene methylation detection kit and method

A detection kit and detection method technology are applied in the field of biomedicine to achieve the effects of high representativeness, low annealing temperature and strong specificity

Pending Publication Date: 2020-01-10
上海中科生物医学高科技开发有限公司
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  • Claims
  • Application Information

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Problems solved by technology

The MSP method is easy to operate and widely used. It can conduct a qualitative study on the free DNA methylation of peripheral blood tumors. It has a good predictive effect on the detection of early malignant tumors, but more sensitive detection methods are still needed in terms of clinical application value.

Method used

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  • Human plasma RASSF1A gene methylation detection kit and method

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Embodiment 1

[0060] 1. Plasma collection

[0061] 1.1 Use a needle above 21G and K3EDTA or K2EDTA vacuum blood collection tube (purple cap) for blood collection, a total of 10ml.

[0062] 1.2 After the blood sample is collected, gently invert the blood collection tube for 5-6 times to ensure that the anticoagulant is fully mixed with the blood;

[0063] 1.3 Place the blood sample tube in the tube rack of the blood collection tube centrifuge and centrifuge at 1,600g for 10min;

[0064] 1.4 After the blood sample is centrifuged, use a filter tip to suck up the plasma 5mm above the white blood cell layer into a 1.5ml EP tube, centrifuge the plasma again at 3,000g for 2min, and suck it into a new EP tube for use.

[0065] 2. DNA extraction from plasma

[0066] 2.1 In a 15ml centrifuge tube, add 3ml of plasma obtained in step 1, 4.5ml of lysate GHH, 300μl of Proteinase K and 1μl of Carrier RNA in sequence, mix well and add 45μl of magnetic beads E.

[0067] 2.2 Vortex to mix, incubate at roo...

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Abstract

The invention provides a human plasma RASSF1A gene methylation detection kit and method. The human plasma RASSF1A gene methylation detection kit includes methylation-specific primer pairs which are designed regarding CpG enrichment sites in human RASSF1A gene promoter regions and have sequences shown in SEQ ID No.1 and SEQ ID No.2. By adopting the human plasma RASSF1A gene methylation detection method, the defects are overcome that a trauma possibility of a surgical specimen during acquiring is high, and a methylation detection rate is relatively low. When detection is conducted regarding theCpG enrichment sites in the RASSF1A gene promoter regions, the specificity is high, acquiring is easy to carry out, and trauma possibilities are low.

Description

technical field [0001] The invention belongs to the technical field of biomedicine, and relates to a human plasma RASSF1A gene methylation detection kit and detection method, in particular to detection in human plasma tumor circulating DNA. Background technique [0002] In recent years, with the in-depth study of the formation mechanism of tumor epigenetics, DNA methylation, as a new molecular marker, has received increasing attention in the diagnosis of tumors. DNA methylation refers to the process in which organisms use s-adenosylmethionine (SAM) as a donor to transfer a methyl group to a base to be determined by DNA methyltransferase (DNAmethyltransferase, DMT). In tumorigenesis, DNA methylation is one of the most common gene abnormalities that can be identified, and the hypermethylation of the promoter region of most specific genes occurs in the early stage of tumor development, and its disorder can be manifested as the overall genome Hypomethylation (hypomethylation) a...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/6886C12N15/11
CPCC12Q1/6886C12Q1/686C12Q2600/154C12Q2563/107C12Q2545/114
Inventor 赵晨阳杨金波张斌
Owner 上海中科生物医学高科技开发有限公司
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