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Digital PCR chip and using method thereof

A chip and digital technology, applied in the field of devices for dispersing trace nucleic acids or cell solutions, can solve the problems of expensive chip detection instruments, difficult to accurately control the sample adding process, etc., and achieve the effects of short sample preparation time, novel design and easy use.

Active Publication Date: 2016-05-04
SUZHOU DIYINAN BIOTECHNOLOGY CO LTD
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  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0003] The problem with microwell PCR-based chips currently on the market is that it is difficult to accurately control the sample addition process, and requires professional training and instrument assistance to achieve the ideal sample dispersion effect.
Moreover, the price of the chip itself and the supporting detection instruments as consumables are also very expensive.

Method used

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  • Digital PCR chip and using method thereof
  • Digital PCR chip and using method thereof
  • Digital PCR chip and using method thereof

Examples

Experimental program
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Effect test

example 1

[0043] Example 1 Dispersing colored dyes with PCR chips

[0044] Step 1: Inject 15 μL of dissolved colored dye into the sample hole of the PCR chip, and under the push of external pressure, flow through the silicon microporous chip, and the hydrophilic dye is dispersed and enters the silicon microporous chip. Due to the hydrophilic-hydrophobic structure of the siliceous microporous silicon wafer, the liquid forms jagged edges during the diffusion process (see Figure 6 ).

[0045] Step 2: Inject sealing oil to fill the cavity outside the silicon microporous chip, and at the same time discharge the air in the cavity to form 20,000 mutually independent closed systems. In this way, the added 15 μL colored dye solution is evenly dispersed into approximately 20,000 microsystems with a volume of approximately 1 nL, indicating that the PCR chip has the potential to uniformly disperse nucleic acid or cell sample solutions (see Figure 5 (a), Figure 5 (b), Figure 10 ).

example 2

[0046] Example 2 Disperse a certain concentration of cell solution with a PCR chip

[0047] Step 1: Dissolve 15 µL at a concentration of 10 6 Each / mL MCF-7 cells were stained with life-and-death staining and DAPI staining respectively, and then injected into the sample wells of the PCR chip respectively. Pushed by external pressure, they flowed through the silicon microporous chip, and the cells in the solution dispersed and entered into the silicon microporous chip. hole chip.

[0048] Step 2: Inject sealing oil to fill the cavity outside the silicon microporous chip, and at the same time discharge the air in the cavity to form 20,000 mutually independent single-cell arrays. Not only can independent cells be clearly observed under bright field (see Figure 7b , Figure 7c ), it is also possible to observe the results of cell life and death staining (see Figure 7a ) and DAPI staining results (see Figure 8a and Figure 8b ). It shows that the PCR chip has the ability t...

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Abstract

The invention provides a digital PCR chip and a using method thereof. The chip comprises a silicon microporous chip, a microfluidic channel for injecting a sample and an aluminum heat conduction base, wherein 20,000 through holes with hydrophilic inner walls and hydrophobic surfaces are processed on the silicon microporous chip. A hydrophilic nucleic acid or cell sample, when flowing through the part above the silicon microporous chip, is spread, monodispersed and retained in each micropore, and mineral oil is introduced, so that the whole space beside the silicon microporous chip is filled, and complete isolation of each microreaction system is realized. The digital PCR chip has the advantages of novel design, easiness in use, low cost and wide application range, and compared with the conventional method for monodispersing the nucleic acid sample in a water-in-oil process, is lowered in cost by more than 10 times, is short in sample preparation time and simple in operation method, is excellent in accuracy of dispersing the nucleic acid sample and reliability of the reaction system, and is small in cross pollution.

Description

technical field [0001] The invention belongs to the technical application field of biochemistry and molecular biology, and in particular relates to a device and a use method for dispersing trace amounts of nucleic acid or cell solution. Background technique [0002] In biological and chemical detection and analysis, polymerase chain reaction (PCR, polymers chain reaction) technology is a powerful research technology and means for nucleic acid. Since KaryMullis invented the PCR method in 1985, PCR instruments have appeared in large and small laboratories all over the world, from the first generation of conventional semi-quantitative end-point PCR technology to the second generation of real-time quantitative PCR technology (Real-timePCR) , and then developed to the current third-generation absolute quantitative digital PCR technology, PCR technology has advanced by leaps and bounds, and is developing in the direction of absolute quantification, high precision, high throughput ...

Claims

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Application Information

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IPC IPC(8): C12M1/00
CPCB01J19/0093B01J2219/00317B01J2219/00013
Inventor 徐峰曹雷崔兴业林敏卢天健
Owner SUZHOU DIYINAN BIOTECHNOLOGY CO LTD
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