Positive transfected cell affinity sorting method and kit

Abstract

The invention relates to a positive transfected cell affinity sorting kit and method. In order to overcome the problem of low gene delivery efficiency in cells difficult to transfect, the invention provides an affinity cell sorting system which can effectively sort transfected positive cells. A series of cell affinity purification tags capable of targeting and positioning the outer surface of a cell membrane are constructed, EGFP fluorescent protein with the affinity tag is expressed in transfection-positive cells and positioned to the surface of the cell membrane under the action of a GPI membrane positioning signal, and on the basis of high-affinity interaction between the affinity tag and a ligand, transfection positive cells are separated through ligand-coupled magnetic beads for detection and quantification. The sorting method is simple to operate, the cells obtained by sorting have small damage, and the method has small interference on gene expression and functions, can be continuously cultured or directly used for downstream experiments, and is especially suitable for solving the problem of low transfection efficiency in cells difficult to transfect.

Description

technical field [0001] The invention belongs to the technical field of sorting positively transfected cells, and specifically relates to an expression cassette, an expression vector containing the expression cassette, a kit, and a method for sorting positively transfected cells based on a fluorescent protein label located on the cell membrane surface and its application . Background technique [0002] The information disclosed in this Background section is only intended to increase the understanding of the general background of the present invention, and is not necessarily to be regarded as an acknowledgment or any form of suggestion that the information constitutes the prior art already known to those skilled in the art. [0003] In recent years, in vitro gene delivery research has made significant progress. High transfection efficiencies can be obtained for most cells, whether viral-mediated or non-viral-mediated gene delivery, however transfection efficiencies in difficu...

AI Technical Summary

Problems solved by technology

However, these molecules themselves have important biological functions, and excessive expression of these molecules on the cell surface may seriously interfere with the gene expression and phenotype of cells, bringing potential unpredictable effects to experimental research

[0005] Therefore, there is still a lack of a method for enriching transfected positive cells that is fast, simple, low in cost, wide in applicability, and less disturbing to cells.

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