High-pressure liquid immersion digital PCR (polymerase chain reaction) method, digital PCR chip and preparation method of digital PCR chip

Abstract

The invention discloses a high-pressure liquid immersion digital PCR (polymerase chain reaction) method, a digital PCR chip and a preparation method of the digital PCR chip. The digital PCR method comprises the following steps: S1, soaking a digital PCR chip after sample feeding treatment into a liquid of a high-pressure reaction chamber; S2, completely discharging air in the high-pressure reaction chamber, and pressurizing the high-pressure reaction chamber after the air is completely discharged; S3, placing the pressurized high-pressure reaction chamber on a PCR instrument, and performing aPCR reaction; S4, cooling the high-pressure reaction chamber after the PCR reaction; S5, reducing the pressure of the cooled high-pressure reaction chamber; and S6, taking out the digital PCR chip inthe high-pressure reaction chamber, and performing fluorescence signal analysis on the digital PCR chip. By adopting the digital PCR method, complex equipment such as pumps and valves is not needed for chip sample feeding, high-viscosity thermal polymerization separation oil is not needed either, the chip does not need to be sealed after sample feeding is completed, and the operation is simple; the chip is small in thickness, rapid in heat conduction and rapid in reaction; and the chip is simple in structure, easy to manufacture, low in cost and high in automation degree.

Description

Technical field [0001] The invention relates to the technical field of nucleic acid detection, in particular to a high-pressure liquid immersion digital PCR method, a digital PCR chip and a preparation method thereof. Background technique [0002] Polymerase chain reaction (Polymerase Chain Reaction, PCR) is a molecular biology technique used to amplify specific DNA fragments. It can be regarded as a special DNA replication in vitro. It has been widely used in molecular biology fields such as genetic testing, gene amplification, genetic engineering, and has played an irreplaceable role in clinical medicine, forensic medicine, paternity testing, and environmental testing. However, the PCR reaction is amplified exponentially, which can be amplified millions of times within tens of minutes, and it is difficult to determine the content of the original PCR template from the PCR product. In order to accurately and quantitatively analyze the nucleic acid content, digital PCR (digital P...

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Problems solved by technology

[0005] After studying the mechanism of bubbles generated by heating the PDMS chip, the present invention proposes a high-pressure liquid immersion digital PCR method, a digital PCR chip and a preparation method thereof, to solve the difficulty and high cost of the digital PCR method in the prior art. The problem with long reaction times

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