Method for rapidly extracting total ribonucleic acid from blood
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A total ribonucleic acid, rapid technology, applied in the field of nucleic acid purification, can solve problems such as cumbersome steps, and achieve the effect of high purity and good integrity
Active Publication Date: 2011-08-17
TIANGEN BIOTECH BEIJING
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Embodiment 1
[0055] Example 1: Extraction of total RNA from human whole blood.
[0056] (1) Add rapid erythrocyte lysate to 400 microliters of human whole blood at a volume ratio of: whole blood: erythrocyte lysate = 1:5, invert and mix 8-10 times to obtain a mixture. Among them, the red blood cell lysate is prepared by mixing 65 grams of ammonium chloride, 18 grams of potassium bicarbonate, 6.5 grams of tetrasodium edetate and 9 grams of cetyltrimethylammonium sulfate, and adding deionized water to obtain the mixed solution, adjust the pH value of the mixed solution to 7.3 with sodium hydroxide solution, and make the mixed solution volume 1000 milliliters.
[0057] (2) Centrifuge the above mixture, the speed of centrifugation is 7300 rpm, the time of centrifugation is 10 minutes, and the centrifuged precipitate is taken out.
[0058] (3) Add 240 microliters of suspension (10 milliliters) to the above precipitate, measure and mix the above components, add deionized water, and make the vol...
Embodiment 2
[0066] Example 2: Extraction of total RNA from whole blood of mice.
[0067] (1) Add rapid erythrocyte lysate to 100 microliters of mouse whole blood at a volume ratio of: whole blood: erythrocyte lysate = 1:5, invert and mix 8-10 times to obtain a mixture. Among them, the red blood cell lysate is prepared by mixing 65 grams of ammonium chloride, 18 grams of potassium bicarbonate, 6.5 grams of tetrasodium edetate and 9 grams of cetyltrimethylammonium sulfate, and adding deionized water to obtain the mixed solution, adjust the pH value of the mixed solution to 7.3 with sodium hydroxide solution, and make the mixed solution volume 1000 milliliters.
[0068] (2) Centrifuge the above-mentioned mixture, the rotating speed of centrifugation is 7300 rpm, the centrifugation time is 3 minutes, and the precipitate of centrifugation is taken out;
[0069] (3) Add 240 microliters of suspension to the above precipitate, grind until the precipitate is completely dissolved, add 200 microlit...
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Abstract
The invention relates to a method for rapidly extracting total ribonucleic acid from blood, belonging to the field of nucleic acid purification. The method comprises the following steps of: firstly, adding a rapid erythrocyte lysing solution into whole blood; performing centrifugal separation on a mixture of the rapid erythrocyte lysing solution and the whole blood; taking precipitate formed by the centrifugal separation out; adding suspension into the precipitate; adding a lysing solution and a protease K solution in sequence; uniformly mixing and then adding absolute ethyl alcohol; uniformly mixing and transferring into a silicon membrane adsorption column to perform adsorptive centrifugation; adding a deproteinizing solution and a deoxyribonuclease solution; performing deproteinizing centrifugation twice; adding a rinsing solution to perform desalting centrifugation; and finally performing drying centrifugation and adding water without ribonuclease, and eluting and centrifuging so as to obtain a ribonucleic acid solution. The method provided by the invention has the characteristics of high efficiency, rapidness and simplicity; and purified RNA (Ribonucleic Acid) can be used for various downstream experiments such as chip analysis, in vitro translation, molecular cloning and the like.
Description
technical field [0001] The invention relates to a method for rapidly extracting total ribonucleic acid from blood, belonging to the field of nucleic acid purification. Background technique [0002] Peripheral blood plays an important role in the immune response and metabolism of humans and animals. As a special tissue, it plays an important role in basic and clinical research because of its easy availability. Peripheral blood can be used as molecular markers in the study of hematopoietic diseases and in the development of in vitro diagnostic systems, and can also be used for molecular monitoring of many systemic diseases outside the blood system. Among them, the most important step in the process of effectively extracting peripheral blood ribonucleic acid (hereinafter referred to as RNA). However, due to the easy coagulation of blood itself, the complexity of various cells in blood, and the fact that it cannot be frozen and then extracted like solid tissue, it is very diffi...
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