Chapter 2
Improved DNA Delivery Efficiency of Bacterial Vectors
by Co-Delivery with Exogenous Lipid and Antimicrobial
Reagents
Andrew N. Osahor and Kumaran Narayanan
Abstract
Gene delivery using invasive bacteria as vectors is a robust method that is feasible for plasmid and artificial
chromosome DNA construct delivery to human cells presenting β1 integrin receptors. This technique is
relatively underutilized owing to the inefficiency of gene transfer to targeted cell populations. Bacterial
vectors must successfully adhere to the cell membrane, internalize into the cytoplasm, undergo lysis, and
deliver DNA to the nucleus. There are limited studies on the use of exogenous reagents to improve the
efficiency of bacteria-mediated gene delivery to mammalian cells. In this chapter, we describe how cationic
lipids, conventionally used for DNA and protein transfection, as well as antimicrobial compounds, can be
used to synergistically enhance the adherence of invasive bacterial vectors to the cell membrane and improve
their predisposition to internalize into the cytoplasm to deliver DNA. Using simple combinatorial methods,
functional DNA transfer can be improved by up to four-fold of invaded cell populations. These methods are
easy to perform and are likely to be applicable for other bacterial vectors including Listeria and Salmonella.
Key words Bactofection, Gene delivery, Invasion, Cationic lipid, Cell penetrating peptide, E. coli,
Transfection efficiency
1 Introduction
Invasive bacteria capable of adherence to the cell membrane and
internalization into the cytoplasm of mammalian cells can be used
as vectors to deliver functional DNA. These vectors are typically
capable of cloning transgenes during their replication cycles and
protecting the DNA during the invasion process [1–3]. These vectors provide a potentially practical approach to circumvent issues
associated with traditional vectors including immunogenicity,
insertional mutagenesis packaging capacity, and production costs
[4–6]. In addition to the delivery of oligonucleotides [1], bacterial
vectors can be used for the in situ production of therapeutic proteins, vaccination, toxin binding, drug delivery, and enzyme
pro-drug therapy [7–9].
Kumaran Narayanan (ed.), Bio-Carrier Vectors: Methods and Protocols, Methods in Molecular Biology, vol. 2211,
https://doi.org/10.1007/978-1-0716-0943-9_2, © Springer Science+Business Media, LLC, part of Springer Nature 2021
15
Improved DNA Delivery Efficiency of Bacterial Vectors
by Co-Delivery with Exogenous Lipid and Antimicrobial
Reagents
Andrew N. Osahor and Kumaran Narayanan
Abstract
Gene delivery using invasive bacteria as vectors is a robust method that is feasible for plasmid and artificial
chromosome DNA construct delivery to human cells presenting β1 integrin receptors. This technique is
relatively underutilized owing to the inefficiency of gene transfer to targeted cell populations. Bacterial
vectors must successfully adhere to the cell membrane, internalize into the cytoplasm, undergo lysis, and
deliver DNA to the nucleus. There are limited studies on the use of exogenous reagents to improve the
efficiency of bacteria-mediated gene delivery to mammalian cells. In this chapter, we describe how cationic
lipids, conventionally used for DNA and protein transfection, as well as antimicrobial compounds, can be
used to synergistically enhance the adherence of invasive bacterial vectors to the cell membrane and improve
their predisposition to internalize into the cytoplasm to deliver DNA. Using simple combinatorial methods,
functional DNA transfer can be improved by up to four-fold of invaded cell populations. These methods are
easy to perform and are likely to be applicable for other bacterial vectors including Listeria and Salmonella.
Key words Bactofection, Gene delivery, Invasion, Cationic lipid, Cell penetrating peptide, E. coli,
Transfection efficiency
1 Introduction
Invasive bacteria capable of adherence to the cell membrane and
internalization into the cytoplasm of mammalian cells can be used
as vectors to deliver functional DNA. These vectors are typically
capable of cloning transgenes during their replication cycles and
protecting the DNA during the invasion process [1–3]. These vectors provide a potentially practical approach to circumvent issues
associated with traditional vectors including immunogenicity,
insertional mutagenesis packaging capacity, and production costs
[4–6]. In addition to the delivery of oligonucleotides [1], bacterial
vectors can be used for the in situ production of therapeutic proteins, vaccination, toxin binding, drug delivery, and enzyme
pro-drug therapy [7–9].
Kumaran Narayanan (ed.), Bio-Carrier Vectors: Methods and Protocols, Methods in Molecular Biology, vol. 2211,
https://doi.org/10.1007/978-1-0716-0943-9_2, © Springer Science+Business Media, LLC, part of Springer Nature 2021
15