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Method for separating ecDNA in exosome based on magnetic beads

A magnetic bead separation and exosome technology, applied in the field of molecular biology, can solve the problems of complex experiments, time-consuming, physical and environmental hazards of operators, etc., and achieve the advantages of simple operation methods, improved experimental efficiency, and reduced experimental manpower Effect

Active Publication Date: 2020-12-25
SHANGHAI CHANGZHENG HOSPITAL +1
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  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0006] However, there are many disadvantages in using cesium chloride gradient centrifugation to extract ecDNA. For example, cesium chloride and ethidium bromide are toxic reagents used in the extraction experiment, which may cause harm to the operator's body and the environment; Many, but also need to use ultracentrifuge, and ecDNA layered absorption technology is difficult, the overall experiment is complicated, and must be operated by trained professionals; the experiment is time-consuming and costly, and cannot be automated

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  • Method for separating ecDNA in exosome based on magnetic beads
  • Method for separating ecDNA in exosome based on magnetic beads
  • Method for separating ecDNA in exosome based on magnetic beads

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Embodiment Construction

[0054] In order to understand the content of the present invention more clearly, it will be described in detail with reference to the drawings and embodiments.

[0055] Such as figure 1 Shown is a schematic flow chart of the method for separating ecDNA in exosomes based on magnetic beads in the present invention, which specifically includes the following steps:

[0056] Exosomes are isolated from biological samples using conventional methods (such as magnetic bead method);

[0057] Using a lysate to lyse the isolated exosomes;

[0058] Use the magnetic bead suspension II to perform nucleic acid adsorption on the products after exosome lysis;

[0059] Washing the magnetic beads adsorbed with nucleic acid using washing solution II with a pH of 7 to 12;

[0060] Using washing solution III containing protease to wash the magnetic beads that have been washed with washing solution II;

[0061] The eluent is used to elute the magnetic beads that have been washed with the washing ...

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Abstract

The invention relates to a method for separating ecDNA in exosome based on magnetic beads. The ecDNA in the exosome is separated in a high-selectivity mode through a magnetic bead method, complex operations such as ultracentrifugation and the like are not needed, and toxic reagents such as cesium chloride and the like are also prevented from being used, so that the method can adapt to automatic operation separation to obtain a relatively high-purity ecDNA product, and the experiment efficiency is greatly improved; and labor and time costs are effectively saved.

Description

technical field [0001] The invention relates to the field of molecular biology technology, in particular to a method for isolating ecDNA in exosomes based on magnetic beads. Background technique [0002] Exosome (exosome) is a 30-150nm size, extremely low-density extracellular nanoscale capsule structure, which is formed by cells through a series of regulatory processes such as "endocytosis-fusion-efflux". Exosomes did not attract the attention of researchers for a long time after they were discovered. Until the introduction of nano-analysis technology and the announcement of the Nobel Prize in Biomedicine in 2013, this particle with a diameter of only tens of nanometers has received unprecedented attention. attention. [0003] Almost all cells secrete exosomes, and the exosomes secreted by cells carry a lot of biological information of the source cells. Therefore, exosomes are also considered as important potential biomarkers, which can reflect the physiological and biolog...

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Application Information

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IPC IPC(8): C12N15/10
CPCC12N15/1013C12Q2521/537C12Q2523/308C12Q2527/125
Inventor 臧远胜马跃张亚楠
Owner SHANGHAI CHANGZHENG HOSPITAL