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Method for purifying non-specific amplification PCR product based on paramagnetic particle method

A non-specific, magnetic bead method, applied in the biological field, to achieve the effect of convenient and fast process and rapid purification

Inactive Publication Date: 2015-04-29
FUJIAN NORMAL UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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There is no report on the method of purifying non-specifically amplified PCR products by using the magnetic bead method

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  • Method for purifying non-specific amplification PCR product based on paramagnetic particle method

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Embodiment 1

[0034] A method for purifying non-specifically amplified PCR products based on the magnetic bead method, characterized in that: it comprises the following steps:

[0035] 1) Detect the PCR product by agarose gel. When the non-specific band is observed to be separated from the target band, cut off the target band with a clean knife, put it into a centrifuge tube, and then add the working band to the centrifuge tube. Solution A, the weight ratio of the target band to working solution A is 1:3, and then incubated at 60°C for 10 minutes;

[0036] 2) Add working solution B equal to the volume of the solution in the above centrifuge tube, shake for 30s, and place at room temperature for 10 minutes, then place the centrifuge tube on the magnetic stand to separate the magnetic beads. When the magnetic beads are adsorbed to the side of the centrifuge tube, Aspirate the liquid and retain the magnetic beads;

[0037] 3) Add 1ml of working solution C to the above-mentioned centrifuge tub...

Embodiment 2

[0044] A method for purifying non-specifically amplified PCR products based on the magnetic bead method, characterized in that: it comprises the following steps:

[0045] 1) Detect the PCR product by agarose gel. When the non-specific band is observed to be separated from the target band, cut off the target band with a clean knife, put it into a centrifuge tube, and then add the working band to the centrifuge tube. Solution A, the weight ratio of the target band to working solution A is 1:3, and then incubated at 50°C for 15 minutes;

[0046] 2) Add working solution B equal to the volume of the solution in the above centrifuge tube, shake for 10s, and place at room temperature for 1min, then place the centrifuge tube on the magnetic stand to separate the magnetic beads. When the magnetic beads are adsorbed to the side of the centrifuge tube, Aspirate the liquid and retain the magnetic beads;

[0047] 3) Add 0.5ml working solution C to the above-mentioned centrifuge tube conta...

Embodiment 3

[0054] A method for purifying non-specifically amplified PCR products based on the magnetic bead method, characterized in that: it comprises the following steps:

[0055] 1) Detect the PCR product by agarose gel. When the non-specific band is observed to be separated from the target band, cut off the target band with a clean knife, put it into a centrifuge tube, and then add the working band to the centrifuge tube. Solution A, the weight ratio of the target band to working solution A is 1:3, and then incubated at 70°C for 8 minutes;

[0056] 2) Add working solution B equal to the volume of the solution in the above centrifuge tube, shake for 20s, and place at room temperature for 5 minutes, then place the centrifuge tube on the magnetic stand to separate the magnetic beads. When the magnetic beads are adsorbed to the side of the centrifuge tube, Aspirate the liquid and retain the magnetic beads;

[0057] 3) Add 1.5ml working solution C to the above-mentioned centrifuge tube c...

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Abstract

The invention discloses a method for purifying a non-specific amplification PCR product based on a paramagnetic particle method. The method comprises the following steps: detecting a PCR product by using agarose gel, when observing that a non-specific strip is separated from a target strip, cutting off the target strip by using a knife, and purifying the non-specific amplification PCR product by using the paramagnetic particle method. The method has the significant advantages that the operation is automatic and the whole process is simple and rapid, and compared with an ordinary purification method, the method is relatively convenient and rapid, and no large-size machine such as a centrifuge is needed in the method, so that relevant study of molecular biology can be relatively well implemented in small-size laboratories.

Description

technical field [0001] The invention belongs to the field of biotechnology, and in particular relates to a method for purifying non-specifically amplified PCR products based on a magnetic bead method. Background technique [0002] Polymerase chain reaction, referred to as PCR. Polymerase chain reaction is a method for enzymatically synthesizing specific DNA fragments in vitro. It consists of several steps such as high-temperature denaturation, low-temperature annealing, and suitable temperature extension. A cycle is carried out, so that the target DNA can be rapidly amplified and has strong specificity. , high sensitivity, easy operation, time-saving and so on. It can be used not only for basic research such as gene isolation, cloning, and nucleic acid sequence analysis, but also for disease diagnosis or wherever there is DNA or RNA. Polymerase chain reaction is also known as cell-free molecular cloning or in vitro primer-directed enzymatic amplification of specific DNA se...

Claims

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Application Information

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IPC IPC(8): C12N15/10
Inventor 王清水余彦
Owner FUJIAN NORMAL UNIV
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