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Kit for rapidly extracting bacterial plasmid DNA by magnetic bead method and extraction method

A bacterial plasmid and kit technology, applied in the field of molecular biology, can solve problems such as time-consuming and poor binding ability

Active Publication Date: 2021-09-14
WUHAN TEXTILE UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0006] However, the above extraction methods or the magnetic beads in the kit have poor binding ability to DNA, and the process of magnetic beads adsorbing DNA takes a long time

Method used

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  • Kit for rapidly extracting bacterial plasmid DNA by magnetic bead method and extraction method
  • Kit for rapidly extracting bacterial plasmid DNA by magnetic bead method and extraction method

Examples

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Effect test

Embodiment 1

[0046] Embodiment 1 provides a kind of kit that magnetic bead method rapidly extracts bacterial plasmid DNA, and said kit includes the lysing reagent group that is used to lyse bacteria and the extraction reagent group that is used to extract plasmid DNA;

[0047] The lysing reagent group consists of bacterial suspension, lysate and neutralizing solution. The resuspension is an aqueous solution containing 15mM ethylenediaminetetraacetic acid and 20mM trishydrochloride, pH 7.5; the lysate is a solution containing 0.3M sodium hydroxide and 2% sodium lauryl sulfate Aqueous solution; Neutralizing solution is an aqueous solution comprising 4M potassium acetate and 3M acetic acid, with a pH of 5.0;

[0048] The extraction reagent group is composed of magnetic bead binding solution, washing solution and eluent. The washing solution is 80% isopropanol aqueous solution or ethanol aqueous solution; the eluent is distilled water.

[0049] The magnetic bead binding solution is a mixed m...

Embodiment 2

[0057] The difference from Example 1 lies in that the preparation ratio of the magnetic bead binding solution is different, and the others are the same as in Example 1, and will not be repeated here.

[0058] In this embodiment, the magnetic bead binding solution is a mixed magnetic bead solution comprising 0.15M sodium acetate, 2wt% dithiothreitol, 3M guanidine isothiocyanate and 3mL silanol magnetic bead suspension with a size of 700nm , wherein the magnetic bead suspension is prepared by mixing magnetic beads and isopropanol at a mass ratio of 1:20, and the pH of the solution is 7.5.

Embodiment 3

[0060] The difference from Example 1 lies in that the preparation ratio of the magnetic bead binding solution is different, and the others are the same as in Example 1, and will not be repeated here.

[0061] In this embodiment, the magnetic bead binding solution is a mixed magnetic bead solution comprising 0.3 sodium acetate, 3wt% dithiothreitol, 3M guanidine isothiocyanate and 3mL silicon carboxyl magnetic bead suspension with a size of 1000nm. The magnetic bead suspension is prepared by mixing magnetic beads and isopropanol at a mass ratio of 1:10, and the pH of the solution is 8.5.

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Abstract

The invention provides a kit for rapidly extracting bacterial plasmid DNA by a magnetic bead method and an extraction method. The kit comprises a lysis reagent group for lysing bacteria and an extraction reagent group for extracting plasmid DNA, wherein the extraction reagent group consists of a magnetic bead binding solution, a washing solution and an eluent. In the method for rapidly extracting plasmid DNA based on the kit, a mixed solution of magnetic beads of which the surfaces are modified with silicon hydroxyl or silicon carboxyl, 0.15-0.45M of sodium acetate, 1-3wt% of dithiothreitol and 3-5M of guanidinium isothiocyanate is taken as a magnetic bead binding solution composite system, so that the binding capacity of plasmid DNA and magnetic beads is enhanced greatly, the extraction efficiency is increased, the extraction speed is increased, the adsorption process of magnetic beads on DNA only needs 1 minute, and extracted DNA has high purity and complete fragments; and meanwhile, reagents are nontoxic, so that environmental pollution is low, and most of similar products on the market can be replaced.

Description

technical field [0001] The invention relates to the technical field of molecular biology, in particular to a kit and an extraction method for rapidly extracting bacterial plasmid DNA by a magnetic bead method. Background technique [0002] In the prior art, the extraction method of bacterial plasmid DNA generally includes steps such as bacterial lysis, DNA aggregation and precipitation, impurity removal, and DNA purification and separation. Traditional extraction methods mainly include column method and CTAB cracking method. These methods are cumbersome and time-consuming, and are not conducive to large-scale extraction. In addition, organic solvents such as chloroform and phenol may be used in the operation process, causing serious pollution. [0003] The magnetic bead method bacterial plasmid DNA extraction kit uses surface-modified magnetic beads and specific adsorption of DNA molecules, and adopts magnetic nano-separation technology, which has the advantages of convenien...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/10
CPCC12N15/1013C12Q2527/125
Inventor 王栋钟亚平鲁振坦
Owner WUHAN TEXTILE UNIV
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