Buffer solution for preparing competent cells and method for preparing competent cells

Abstract

The invention discloses a buffer solution for preparing competent cells and a method for preparing the competent cells, belonging to the field of biotechnologies. The buffer solution is an aqueous solution containing piperazine-1,4-bisethanesulfonic acid, N,N-(2-hydroxyethyl)-2-aminoethanesulfonic acid, calcium chloride, potassium chloride and manganese chloride, wherein the pH value of the buffer solution is 6.0-7.0, and the concentrations of piperazine-1,4-bisethanesulfonic acid, N,N-(2-hydroxyethyl)-2-aminoethanesulfonic acid, calcium chloride, potassium chloride and manganese chloride are respectively 5-30 mM, 3-20 mM, 5-25 mM, 50-300 mM and 10-50 mM. For the buffer solution provided by the embodiment of the invention, by adding N,N-(2-hydroxyethyl)-2-aminoethanesulfonic acid, the permeability of the cell membrane can be increased, thus the conversion efficiency of the prepared competent cells is greatly improved, meanwhile, the prepared competent cells are suitable for transformation of long fragments, and moreover, with the preparation method provided by the embodiment of the invention, the competent cells can be efficiently and rapidly prepared.

Description

technical field [0001] The invention relates to the field of biotechnology, in particular to a buffer for preparing competent cells and a method for preparing competent cells. Background technique [0002] The method of transforming recipient bacteria with exogenous DNA is one of the most commonly used methods in molecular biology research, and exogenous DNA includes plasmids and linking vectors. Exogenous DNA must be transferred into appropriate recipient bacteria for propagation in order to obtain a large number of positive recombinant clones or express the target gene product. [0003] The existing method is to culture the recipient bacteria in LB plate medium, pick a single colony and inoculate it in SOB (SuperOptimal Broth) shake flask medium for fermentation to realize the reproduction of a single colony, and shake culture at 18°C ​​to OD 600 Value is 0.6, ice bath for 10min, centrifuge at 4°C for 10min, bacteria suspended in pre-cooled TB buffer, ice-bathed for 10min...

AI Technical Summary

Problems solved by technology

[0004] In order to solve the problem of low transformation efficiency of the preparation of competent cells by chemical methods in the prior art, the embodiment of the present invention provides a buffer for preparing competent cells and a method for preparing competent cells

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