pH-sensitive nucleic acid quantitative detection method and device

Abstract

The invention discloses a pH-sensitive nucleic acid quantitative detection method and device. The method comprises the following steps: S1, performing a nucleic acid amplification reaction by using a plurality of reference templates, and detecting the real-time pH value of an amplification reaction solution; S2, obtaining an S-shaped curve of the pH value of each reference template along with time change in the amplification process, and determining the pH threshold value of the S-shaped curve of each reference template in the rapid amplification period; S3, establishing an initial concentration-time standard curve according to the initial concentration of each reference template and the time for reaching the pH threshold in the amplification process; S4, performing nucleic acid amplification reaction on a to-be-detected sample under the same condition as that in the S1, detecting the real-time pH value, and determining the time for reaching the pH threshold value, wherein the nucleic acid types and sequences of the to-be-detected sample and the reference template are the same; and S5, determining the initial concentration of the to-be-detected sample according to the initial concentration-time standard curve and the time determined in the step S4. According to the invention, the structure simplification and miniaturization of the detection device can be realized, the operation is convenient, and the detection sensitivity and accuracy are high.

Description

technical field [0001] The invention relates to the field of nucleic acid detection, in particular to a pH-sensitive nucleic acid quantitative detection method and device. Background technique [0002] Nucleic acid quantitative detection is a technology that amplifies nucleic acid and monitors the amplification process by different means to obtain the initial concentration of nucleic acid before amplification. Ordinary polymerase chain reaction (PCR) can only roughly detect the amount of amplified product after the reaction is over, the so-called endpoint method, which cannot be used for quantitative detection of nucleic acid in the sample. This is mainly because, as the reaction proceeds, primers and nucleotide starting materials are continuously consumed, and ultimately cannot be exponentially amplified. Therefore, in traditional PCR, there is no linear relationship between the final yield of amplified products and the initial copy number of the template. [0003] To rea...

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However, the method of fluorescence quantification requires an optical path device for excitation and detection of fluo

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