Biomedical Engineering Reference
In-Depth Information
a
Wild type virus genome
packaging
signal
Essential viral genes
Ψ
b
Viral vector DNA
Viral
regulatory
sequence
Essential viral genes
packaging
signal
transgene
Ψ
c
Helper DNA (
ΔΨ
)
Viral
regulatory
sequence
Transfection
Integration
of helper DNA
Transfection of viral
vector DNA
into packaging cell
e
Producer Cell
Production of viral
replication and
structural proteins
Viral particles
containing
transgene
Packaging cell
d
Purification of
virus particles
and infection of
target cells
f
Viral RNA/DNA
intermediates
(integration into
host cell genome)
g
transcription
and translation
Target Cell
h
Protein of interest
Figure 11.1
Strategy for viral vector production. (Modified from Lawson [17].) (a) Typical
viral genome with essential viral genes, regulatory sequences and packaging signal. In order to
produce replication-deficient viral vectors the genetic material is separated onto (b) plasmid
encoding viral packaging signal and minimum viral regulatory sequences and (c) helper plasmids
encoding essential viral genes required for viral replication and packaging. (d) Production of
packaging cell line by transfection of helper plasmids. (e) Production of producer cell line by
transfection of packaging cell line with viral vector plasmid. (f) Isolation and purification of
virus particles from producer cell culture supernatants (e.g. retrovirus vectors) or lysates (e.g.
Ad5 vectors). (g) Infection of target cells with viral vectors and (h) expression of protein of
interest in target cells (can be secreted, cell surface or intracellular protein, or could be siRNA
to inhibit expression of endogenous protein).
