Nucleic acid extraction composition, reagent and kit containing the same and use thereof
A nucleic acid extraction composition, reagents and kits containing the same and uses thereof. Provided is a nucleic acid extraction and purification reagent free of volatile organic solvents, which prevents the damage of volatile organic solvents to people and greatly improves the timeliness of nucleic acid extraction and purification, making the operation extremely simple, and the nucleic acid can be obtained within 10 minutes. The obtained nucleic acid may be used for biological reactions such as PCR, NASBA, LAMP and RPA. Moreover, the reagents of the present disclosure may be used to extract nucleic acids of cells, bacteria, fungi, DNA viruses and RNA viruses from various samples such as blood, throat swab preserving fluid, saliva, urine, sputum, excrement and the like, and very suitable for clinical and scientific research uses.
1. A nucleic acid extraction composition comprising a binding buffer, a wash buffer, a lysis buffer, an enhancer, and an elution buffer, wherein
the binding buffer is a composition comprising glycogen, 3-(Morpholin-4-yl)propane-1-sulfonic acid (MOPS), polyamine, and tris(2-carboxyethyl)phosphine (TCEP); the wash buffer is a composition comprising guanidine hydrochloride, potassium acetate, glycogen, MOPS, and polyamine;
in the binding buffer, the concentration of glycogen is 0.1% to 1% (m/v), the concentration of MOPS is 10 mM to 20 mM, the concentration of polyamine is 0.1% to 2% (m/v), and the concentration of TCEP is 5 mM to 50 mM; the polyamine comprises spermine, spermidine, butanediamine or a mixture thereof; and the pH of the binding buffer is 8.5 to 10;
in the wash buffer, the concentration of guanidine hydrochloride is 0.05 M to 0.6 M, the concentration of potassium acetate is 0.05 M to 0.5 M, the concentration of glycogen is 0.01% to 0.1% (m/v), the concentration of MOPS is 1 mM to 2 mM, and the concentration of polyamine is 0.01% to 0.3% (m/v); the polyamine comprises spermine, spermidine, butanediamine or a mixture thereof; and the pH of the wash buffer is 5 to 6;
the lysis buffer comprises guanidine isothiocyanate, 2-[4-(2,4,4-trimethylpentan-2-yl)phenoxy]ethanol or a mixture thereof; and in the lysis buffer, the concentration of guanidine isothiocyanate is 2 M to 8 M, the concentration of 2-[4-(2,4,4-trimethylpentan-2-yl)phenoxy]ethanol is 5% to 20% (v/v), and the pH value is 3 to 6;
the enhancer comprises proteinase K, trehalose, calcium chloride or a mixture thereof; and in the enhancer, the concentration of proteinase K is 10 mg/ml to 25 mg/ml, the concentration of trehalose is 3% to 12% (m/v), and the concentration of calcium chloride is 5 mM to 20 mM;
the elution buffer comprises sodium hydroxide, 1×Tris-EDTA (TE) buffer or a mixture thereof; the pH of 1×TE buffer in the elution buffer is adjusted to 8 to 10 with sodium hydroxide; and
the nucleic acid extraction composition does not comprise volatile organic solvents.
2. A kit for nucleic acid extraction or nucleic acid detection comprising the nucleic acid extraction composition according to claim 1 and a reagent or carrier acceptable for extraction and/or detection.
3. The kit according to claim 2 , wherein the carrier is selected from the group consisting of a silicon-based adsorption column, magnetic beads, glass beads, and a chitosan or silica modified carrier.
4. A method for nucleic acid extraction or detection for non-diagnostic purposes comprising
mixing a nucleic acid sample to be tested with the nucleic acid extraction composition according to claim 1 ; and
performing extraction and/or detection.