Plasmid production and purification: An integrated experiment‐based biochemistry and biotechnology laboratory course

Abstract

This laboratory experiment describes the production and purification of plasmid DNA for undergraduate biochemistry and biotechnology courses. This experiment performed in a one‐week period includes the protocols for plasmid pVAX1‐LacZ production in Escherichia coli DH5α cells and subsequent purification of supercoiled pVAX1‐LacZ. Firstly, the students use a growth medium that favors the replication of the plasmid resulting in a higher plasmid production during exponential growth. Afterwards, alkaline lysis is done to disrupt the bacterial cells and recover pVAX1‐LacZ plasmid. Lastly, they perform the purification of pVAX1‐LacZ supercoiled isoform by L‐histidine chromatography, followed by agarose gel electrophoresis to characterize the separation of supercoiled isoform from contaminants. The proposed experiment provides an opportunity for students to acquire these skills that are routinely used in biochemistry and biotechnology laboratories. © 2019 International Union of Biochemistry and Molecular Biology, 47(6):638–643, 2019.

Nuestros microcompañeros

“…Una característica descubierta en algunos microorganismos es su capacidad para intercambiar secciones de su secuencia de ADN; a estos fragmentos de ADN dinámicos se les denomina transposones (Tn) que son los responsables de la rápida respuesta de los microorganismos hacia los antibióticos (Bustos et al, 2017;Santos et al, 2019). El hecho de que los Tn se sitúen en un plásmido contribuye a que la resistencia a los antibióticos se transfiere entre bacterias distintas (Bustos et al, 2017).…”