Nucleic acid detection micro-fluidic chip and preparation method thereof

Abstract

The invention relates to a nucleic acid detection micro-fluidic chip, which comprises a substrate and a cover plate, wherein the cover plate is fixedly arranged on the substrate; the cover plate is provided with an oil phase sample-injection hole and a sample sample-injection hole; the surface, abutted with the substrate, of the cover plate is a structural surface; a first micro-flow channel connected with the oil phase sample-injection hole, a second micro-flow channel connected with the sample sample-injection hole and a liquid drop generation interface located in the joint of the first andsecond micro-flow channels, which are mutually communicated, are etched in the structural surface; the structural surface, in a liquid drop collection area, of the cover plate is fixedly provided witha thin plate; a gap is formed between the periphery of the thin plate and the boundary of the liquid drop collection area, so as to form a gas path; and the cover plate is also provided with a gas vent hole communicated with the gas path. The problems of the fusion and the demulsification of liquid drops are avoided; the homogeneity and the stability of the liquid drop are improved; meanwhile, the signal reading speed of the liquid drop is quickened during dPCR (digital Polymerase Chain Reaction) detection; and the detection period is shortened. The invention also relates to a preparation method of the nucleic acid detection micro-fluidic chip. The preparation method is simple and convenient; the cost is low; and the nucleic acid detection micro-fluidic chip not only can be produced in alarge-batch manner, but also can be produced in a laboratory in a small-batch manner.

Description

technical field [0001] The invention belongs to the field of nucleic acid detection, and in particular relates to a nucleic acid detection microfluidic chip and a preparation method thereof. Background technique [0002] Since the introduction of polymerase chain reaction (polymerase chain reaction, PCR), in vitro nucleic acid amplification and quantification technology has developed into a core technology in the field of molecular biology. At present, PCR-based nucleic acid quantitative detection technology has become the main application technology for early diagnosis of diseases due to its remarkable advantages of fast analysis speed, high sensitivity, high throughput and short diagnosis time, and is widely used in molecular sequencing, gene expression Other research fields such as analysis, gene mutation study and drug screening. [0003] ABI Corporation of the United States has launched real-time fluorescent quantitative PCR (Real-time quantitative PCR, qPCR) technolog...

AI Technical Summary

Problems solved by technology

The system can disperse the droplets into millions of droplets. However, the system uses flow analysis technology to analyze millions of droplets one by one in the flow cytometer, and the signal reading takes a long time, resulting in a detection cycle Long, the entire detection process can last up to 3.5 hours

In addition, since the droplets need to be transferred to the flow cytometer for analysis, it is difficult to avoid contamination and demulsification during the transfer process

In addition, when the system generates droplets and collects them for PCR reaction, they are not arranged in a single layer of droplets, but are collected into a sample tank, which will cause some droplets to fuse, resulting in low uniformity of droplets in the system

[0008] The continuous phase (mainly composed of mineral oil, CAS: 8042-47-5) that is currently used to generate droplets and is compatible with the reaction conditions of common nucleic acid detection reactions PCR, and the microfluidic chip commonly used to produce droplets The material PDMS is not compatible, PDMS will absorb mineral oil, which is not obvious at room temperature, but as the temperature rises, PDMS absorbs mineral oil faster, and will be quickly absorbed at a high temperature of 95°C, resulting in droplets due to continuous phase This is the difficulty in realizing the liquid droplet digital PCR reaction in the chip

[0009] In addition, due to the droplet digital PCR technology in the process of using the chip, there are two immiscible fluids, namely the dispersed phase and the continuous phase. The dispersed phase and the continuous phase intersect at the droplet generation interface, and the dispersed phase is sheared into a A mutually independent spherical droplet is dispersed in the continuous phase, and the continuous phase remains connected as a whole; when the chip starts to work, due to the great difference in the properties of the two-phase fluids, the two-phase fluids usually do not reach the droplet generation interface at the same time. Bubbles will be mixed in when the droplets are produced, and the bubbles will affect the preservation of the droplets. That is, if the bubbles are mixed into the droplet collection area, due to the heating process in the reaction process, the bubbles will expand rapidly due to heating, making the storage environment of the droplets Instability, which affects the stability of the droplet itself, will also cause the droplets to fuse together, and even push the droplets out of the droplet collection area

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