The invention discloses a method for fast and accurate
quantitative determination of
circulating DNA in blood. The method comprises the following specific steps that
processing of
plasma or serum is performed:
peripheral venous blood is taken and arranged in an EDTA
anticoagulant tube for standing at
room temperature, or self-clotting
whole blood is taken to undergo
centrifugation under 1800-2200 g of
centrifugal force for 4 to 6 minutes, upper-layer
plasma is taken, the same volume of diluted
reagent is added, even mixing is performed, stirring is performed at the temperature of 92-97 DEG C for 6 to 10 minutes, then
centrifugation is performed again under 15000-17000 g of
centrifugal force for 8 to 12 minutes, and supernate is taken as a template for PCR; a quantitative PCR
reaction system is configured; quantitative PCR reaction is performed. The quantitative PCR reaction can be performed by directly using the processed
plasma or serum, PCR reaction is not influenced, the amplification efficiency is consistent with the amplification efficiency of purified
DNA,
DNA extraction steps and experimental errors are decreased, a new fluorescent dye SuperGreen which is higher in concentration, high in fluorescent
signal value and
response sensitivity and capable of accurately detecting plasma
free DNA as low as the nanogram level can be adopted, the
reagent price is low, the using cost of
a DNA extraction kit is reduced, and operation is easy and convenient.