Single nucleotide polymorphism analysis method based on magnetic functionalized microfluidic chip

A single nucleotide polymorphism, microfluidic chip technology, applied in biochemical equipment and methods, microbial determination/inspection, etc., can solve the problem that the single nucleotide polymorphism analysis method has not been reported, and achieve Good biocompatibility and hydrophilicity, maintaining activity, efficient separation and detection

Active Publication Date: 2018-12-18
NANCHANG UNIV
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  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

However based on GO@Fe 3 o 4 SNP analysis methods for...

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  • Single nucleotide polymorphism analysis method based on magnetic functionalized microfluidic chip
  • Single nucleotide polymorphism analysis method based on magnetic functionalized microfluidic chip
  • Single nucleotide polymorphism analysis method based on magnetic functionalized microfluidic chip

Examples

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Embodiment 1

[0028] (1) Preparation of graphene oxide: 0.5g graphite powder and 0.5gNaNO 3 Added to 23 mL of concentrated H 2 SO 4 In the ice bath, slowly add 3gKMnO 4 , stir well to make it evenly mixed, transfer the solution to a water bath at 35°C and stir for 1 hour to form a beige paste, add 40mL of ultrapure water, stir at room temperature for 30min, add ultrapure water to dilute to 140mL, drop by drop Add 3 mL of 30% HO 2 o 2 , the solution changed from dark brown to bright yellow, and the obtained product was filtered while hot, and the product was centrifuged with ultrapure water until the supernatant was neutral, and centrifuged at 8000r / min for 2min to obtain graphene oxide (GO).

[0029] (2) Preparation of GO@Fe 3 o 4 Nanocomposite material: Dissolve 40mg GO in 20mL ultrapure water, ultrasonically disperse for 3h, and then pass N 2 Under the condition of heating to 70 ℃ and keeping it for 30min, then add 21mg FeCl 3 ·6H 2 O and 72 mg FeCl 2 4H 2 O, stirred for 40min;...

Embodiment 2

[0033] Preparation of GO@Fe 3 o 4 Functionalized PDMS microfluidic chip separation channel: first rinse the PDMS microfluidic separation channel with ultrapure water for 5 minutes, then place a permanent magnet with a length of 2 cm on the top and bottom of the chip, and vacuum the GO prepared in Example 1 @Fe 3 o 4 The solution is pumped into the separation channel, GO@Fe 3 o 4 It was immobilized in the PDMS microfluidic separation channel, and the modified microfluidic chip was placed for 1 hour, and the separation channel was continuously washed with running buffer solution for 5 minutes to form GO@Fe 3 o 4 Functionalized PDMS microfluidic chip separation channel.

[0034] GO@Fe will be produced 3 o 4 The functionalized PDMS microfluidic chip was used to characterize the contact angle, and the PDMS bare chip and GO-modified PDMS chip were used as controls. The results are as follows: image 3 shown. Depend on image 3 A shows that the contact angle of the PDMS mi...

Embodiment 3

[0037] GO@Fe 3 o 4 Analysis of Single Nucleotide Polymorphism by Functionalized PDMS Microfluidic Chip

[0038]Design a methylene blue-labeled probe DNA (P1, 5'-methylene blue-TCAACATCAGTCTGATAAGCTA-3') that is completely complementary to mircoRNA-21 (T1, 5′-UAGCUUAUCAGACUGAUGUUGA-3′), P1 hybridizes with T1 to form a completely complementary double strand (P1T1), P1 hybridizes with the single-base mismatch strand sm-mircoRNA-21 (M1, 5′-UAGCUUAUAAGACUGAUGUUGA-3′) of T1 to form a single-base mismatch double strand (P1M1). Mix different concentrations of T1, 50 μL of 2.5 μM P1 and 20 μL of hybridization buffer solution (20 mM Tris-HCl buffer solution, pH 7.4), add ultrapure water to make the final volume of the solution 200 μL, and perform hybridization reaction at 37°C for 2.5 h. The hybridization reaction between M1 and P1 was carried out in the same steps as above. The hybridization reaction between different concentrations of T1 and different concentrations of M1 and 0.5 μ...

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Abstract

The invention discloses a single nucleotide polymorphism analysis method based on a magnetic functionalized microfluidic chip, which belongs to the technical field of the microfluidic chip. The methodcomprises the following steps: Fe3O4 magnetic nanoparticles are loaded on the surface of graphene oxide to synthesize magnetic nanocomposites (GO@Fe3O4), under the action of an external magnetic field, GO@Fe3O4 is efficiently and controllably fixed in a separation channel of the microfluidic chip to obtain a GO@Fe3O4 functionalized microfluidic chip separation channel. In addition, the method successfully implements the analysis of single nucleotide polymorphism in the GO@Fe3O4 functionalized microfluidic chip separation channel, and the analysis method has the advantages of simplicity, convenience, high efficiency and short time.

Description

technical field [0001] The invention relates to a single nucleotide polymorphism analysis method based on a magnetically functionalized microfluidic chip, belonging to the technical field of microfluidic chips. Background technique [0002] Single nucleotide polymorphisms (SNPs) are single base variations in genetic loci that are closely related to various human genetic diseases. It is the most common type of heritable variation in humans. MicroRNAs (miRNAs) are a group of endogenous small non-coding RNAs with a length of 18-25 nucleotides. As an important post-transcriptional regulator, miRNA is involved in various developmental and physiological processes of animals and plants, and also plays an important role in diseases. MiRNAs can act as oncogenes or tumor suppressors, regulating the expression of about one-third of human genes. MiRNAs affect cancer risk, and mutations in related genes will affect the function of miRNAs, thereby affecting a variety of biological proc...

Claims

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Application Information

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IPC IPC(8): C12Q1/6858
CPCC12Q1/6858C12Q2565/629C12Q2563/137
Inventor 梁汝萍吴露露邱建丁
Owner NANCHANG UNIV
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