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Integrated digital PCR system and using method thereof

A digital and fluid technology, applied in the field of integrated digital PCR system, can solve the problems of complex use process, insufficient reliability, and high operating cost, and achieve the effects of simple operation, avoiding heating and cooling processes, and low cost.

Pending Publication Date: 2020-11-10
CHONGQING UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] Aiming at the above-mentioned deficiencies in the prior art, an integrated digital PCR system and its use method provided by the present invention solve the problems of complex use process, high operation cost, low detection efficiency and insufficient reliability of the existing digital PCR system

Method used

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  • Integrated digital PCR system and using method thereof
  • Integrated digital PCR system and using method thereof
  • Integrated digital PCR system and using method thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0068] Such as figure 1 As shown, an integrated digital PCR system includes a digital PCR chip 1, an optical detection module 2, a fluid control module 3, a hot water circulation temperature control module 4 and a central control module 5;

[0069] The digital PCR chip 1 is placed on the two-dimensional step scanning platform 2-6 on the optical detection module 2, and is connected with the hot water circulation temperature control module 4 and the fluid control module 3, and the fluid control module 3 is also connected with the hot water circulation temperature control module respectively. The control module 4 and the central control module 5 are connected, and the hot water circulation temperature control module 4 and the optical detection module 2 are connected with the central control module 5 .

[0070] Among them, the digital PCR chip 1 is used to realize the automatic decomposition and quantitative uniform distribution of input samples, without the need to configure comp...

Embodiment 2

[0095] Such as Figure 5 As shown, the present embodiment provides a method for using the digital PCR system in the above embodiment 1, including the following steps:

[0096] S1. Preparation: Fix the digital PCR chip on the two-dimensional stepping scanning platform, assemble and connect the through pipes according to the requirements of each interface, start the central control module at the same time, set the parameters for regulating fluid control and temperature control, and start the temperature The control circuit controls the temperature of the constant temperature water tank in the hot water circulation temperature control module according to the detection requirements of digital PCR;

[0097] S2. Sample filling: adjust the opening state of the valve based on the detection requirements of digital PCR, and use a vacuum pump to extract the air in each micropipe and microcavity in the digital PCR chip, and then suck the sample liquid in the sample tube into the digital P...

Embodiment 3

[0121] Such as Figure 6 As shown, using the system of the present invention to perform conventional digital PCR detection, taking the detection of EGFRL858R mutation in the peripheral blood of a lung cancer patient as an example, the specific steps are as follows:

[0122] (1) Chip, reagent and sample preparation

[0123] An integrated cavity digital PCR chip containing 20,000 microcavities was fabricated by photolithography molding process combined with PDMS pouring mold and oxygen plasma surface treatment bonding process; a pair of specific primers were synthesized for EGFR L858R mutation; Circulating DNA is extracted from blood serum, and polymerase, primers, probes, buffers, etc. are added to the sample tube storing circulating DNA, mixed to prepare a sample solution, and paraffin oil is added to the sample solution.

[0124] (2) Chip Assembly

[0125] The digital PCR chip prepared in step (1) is placed and fixed on the two-dimensional stepping scanning platform, and th...

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Abstract

The invention discloses an integrated digital PCR system and a using method thereof. The system involves a high-density micro-cavity array digital PCR chip integrated with one cavity, an optical detection module, a hot water circulation temperature control module, a fluid control module and a central control module. When the system is applied to nucleic acid sample detection, firstly, the digitalPCR chip is fixed on an objective table of the optical detection module, and through pipes communicating with a sample pipe and a hot water tank are assembled at interfaces of the chip; and then, thecentral control module is started, relevant parameters of a control program are set, and the fluid control module, the hot water circulation temperature control module and the optical detection moduleare controlled through the control program to sequentially achieve sample filling, sample discretization, hot water circulation amplification, chip scanning recording, data analysis and other operations. Based on the combination of the micro-cavity array and the integrated cavity structure of the digital PCR chip, the system can enable the whole process of digital PCR detection to be completed ona single instrument, and the detection efficiency and the result reliability are improved.

Description

technical field [0001] The invention belongs to the technical field of microfluidic chip analysis, and in particular relates to an integrated digital PCR system and a using method thereof. Background technique [0002] Digital PCR (digital PCR, dPCR) technology is the third generation of quantitative nucleic acid detection technology following the real-time fluorescent quantitative PCR (Quantitative Real-time PCR) technology. It is an absolute quantitative technology of nucleic acid. Compared with traditional PCR, it has the following advantages Advantages: It can achieve absolute nucleic acid quantification without relying on the standard curve, has high tolerance to inhibitors, can analyze complex mixtures, can detect trace nucleic acid samples, detect rare mutations in complex backgrounds, and identify small differences in expression levels, etc. It works by decomposing the sample into tens of thousands or even millions of droplets (each droplet acts as an independent PCR...

Claims

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Application Information

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IPC IPC(8): C12M1/38C12M1/36C12M1/34C12M1/00C12Q1/6851
CPCB01L3/502784B01L7/52B01L2300/0861B01L2300/185B01L2400/0487C12Q1/6851C12Q2563/107C12Q2563/159C12Q2565/629
Inventor 李刚谢腾宝武银
Owner CHONGQING UNIV
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