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Silica gel absorption column extraction kit and method for free DNA (Deoxyribonucleic Acid) and RNA (Ribonucleic Acid) of blood plasma or blood serum
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A technology of silica gel adsorption and kits, which is applied in the biological field and can solve problems such as high prices
Inactive Publication Date: 2018-05-25
江苏然科生物技术有限公司
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At present, the most important product of the silica gel membrane adsorption column method is QIAamp Circulating Nucleic Acid Kit from Qiagen, which is very expensive because it has a very large monopoly in the field of free DNA or RNA extraction.
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[0104] Wherein, the substance concentration of Tris-HCl can realize the technical purpose of the present invention at 10-100mM, for example, it can be: 10mM, 20mM, 30mM, 40mM, 60mM, 70mM, 80mM, 90mM, etc.;
[0105] The amount concentration of guanidineisothiocyanate can realize the technical purpose of the present invention at 3-5M, for example can be: 3M, 3.5M, 4M, 4.5M, 5M etc.;
[0106] The substance concentration of sodiumchloride can realize the technical purpose of the present invention at 0.5-1.5M, for example can be: 0.5M, 0.7M, 0.9M, 1.1M, 1.3M, 1.5M etc.;
[0130] Take fresh EDTA anticoagulated blood, centrifuge at 900G for 10 minutes, and absorb the supernatant; centrifuge the supernatant at 15,800G for 10 minutes, absorb the supernatant to obtain plasma, divide the prepared plasma into 4 parts on average, and set aside.
[0131] Among them, 2 parts of the mixed plasma were extracted with the method and kit of the present invention, and the remaining 2 parts of the mixed plasma were extracted with Qiagen's QIAamp Circulating Nucleic Acid Kit to extract plasma free DNA and RNA.
[0133] Add an equal volume of lysate to 4ml plasma, add 320μl proteinase K, mix for 30s, and incubate at 60°C for 30 minutes.
[0134] Wherein, the volume ratio of the amount of plasma, lysate and proteinase K can realize the technical purpose of the present invention in the scope of 0.5-1.5:0.5-1....
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technical field [0001] The invention relates to the field of biotechnology, in particular to a kit and method for extracting plasma or serum freeDNA and RNA through a silica gel adsorption column. technical background [0002] Free DNA, referred to as cf DNA (Free circulating / Cell-free DNA), was first discovered in 1948. It is a cell-free state, fragmented extracellular DNA, and mainly exists in body fluids such as blood, synovial fluid, and cerebrospinal fluid. At present, the application of cell-free DNA in non-invasive prenatal detection and liquid biopsy of tumors are two hot fields in clinical practice. In addition, the application value of cell-free DNA in diseases such as organ transplantation, stroke, autoimmune disease and myocardial infarction has become increasingly clear. [0003] Free RNA (cell-freeRNA, cfRNA) is the transcription product existing outside the cell, including mRNA and microRNA. MicroRNA is found in a variety of human body fluids (plasma, serum...
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