Chapter 12
Electroporation of Mycobacteria
Tanya Parish
Abstract
The introduction of DNA into bacterial cells is one of the foundational methods of bacterial genetics.
Transformation of mycobacterial species is complicated due to the structure of the cell wall, which has a
complex outer layer with low permeability. Electroporation has become a routine procedure in genetic
studies. In this process, cells are subjected to a brief high-voltage electrical impulse which allows the entry of
DNA. It can be used to introduce plasmid DNA, phage DNA, or oligonucleotides. This chapter presents
methods for introducing DNA into a representative slow-growing species, M. tuberculosis, and a representative fast-growing species, M. smegmatis. Other mycobacteria can be transformed using variations of these
methods, although the efficiency of transformation will vary.
Key words Mycobacteria, Electroporation, Bacterial Genetics, Plasmids, DNA delivery
1 Introduction
The introduction of DNA into bacterial cells is one of the foundational methods of bacterial genetics. Mycobacteria are notoriously
difficult bacterial species to manipulate, due to their thick cell wall
and their slow growth rate. The introduction of DNA into these
species is complicated due to the structure of the cell wall, which
has a complex outer layer with low permeability. Electroporation is
one of the key methods to introduce DNA into mycobacteria and
has become a routine procedure in genetic studies. In this process,
cells are subjected to a brief high-voltage electrical impulse which
allows the entry of DNA. It can be used to introduce plasmid DNA,
phage DNA, or oligonucleotides [1].
To date, numerous species of mycobacteria have been successfully transformed with DNA by electroporation including Mycobacterium tuberculosis [2], Mycobacterium bovis BCG [2],
Mycobacterium vaccae [3], Mycobacterium w [4], Mycobacterium
phlei [5], Mycobacterium fortuitum [5], Mycobacterium aurum
[6], Mycobacterium intracellulare [7], Mycobacterium parafortuitum [6], Mycobacterium marinum [8]t, Mycobacterium avium [9],
Tanya Parish and Anuradha Kumar (eds.), Mycobacteria Protocols, Methods in Molecular Biology, vol. 2314,
https://doi.org/10.1007/978-1-0716-1460-0_12, © Springer Science+Business Media, LLC, part of Springer Nature 2021
273
Electroporation of Mycobacteria
Tanya Parish
Abstract
The introduction of DNA into bacterial cells is one of the foundational methods of bacterial genetics.
Transformation of mycobacterial species is complicated due to the structure of the cell wall, which has a
complex outer layer with low permeability. Electroporation has become a routine procedure in genetic
studies. In this process, cells are subjected to a brief high-voltage electrical impulse which allows the entry of
DNA. It can be used to introduce plasmid DNA, phage DNA, or oligonucleotides. This chapter presents
methods for introducing DNA into a representative slow-growing species, M. tuberculosis, and a representative fast-growing species, M. smegmatis. Other mycobacteria can be transformed using variations of these
methods, although the efficiency of transformation will vary.
Key words Mycobacteria, Electroporation, Bacterial Genetics, Plasmids, DNA delivery
1 Introduction
The introduction of DNA into bacterial cells is one of the foundational methods of bacterial genetics. Mycobacteria are notoriously
difficult bacterial species to manipulate, due to their thick cell wall
and their slow growth rate. The introduction of DNA into these
species is complicated due to the structure of the cell wall, which
has a complex outer layer with low permeability. Electroporation is
one of the key methods to introduce DNA into mycobacteria and
has become a routine procedure in genetic studies. In this process,
cells are subjected to a brief high-voltage electrical impulse which
allows the entry of DNA. It can be used to introduce plasmid DNA,
phage DNA, or oligonucleotides [1].
To date, numerous species of mycobacteria have been successfully transformed with DNA by electroporation including Mycobacterium tuberculosis [2], Mycobacterium bovis BCG [2],
Mycobacterium vaccae [3], Mycobacterium w [4], Mycobacterium
phlei [5], Mycobacterium fortuitum [5], Mycobacterium aurum
[6], Mycobacterium intracellulare [7], Mycobacterium parafortuitum [6], Mycobacterium marinum [8]t, Mycobacterium avium [9],
Tanya Parish and Anuradha Kumar (eds.), Mycobacteria Protocols, Methods in Molecular Biology, vol. 2314,
https://doi.org/10.1007/978-1-0716-1460-0_12, © Springer Science+Business Media, LLC, part of Springer Nature 2021
273