Biology Reference
In-Depth Information
Schneemann
, 2006).
Intact VLPs indistinguishable from native particles were self-assembled
and could be isolated in all three cases; FHV VLPs are shown in Fig. 3.5.
Compared with other expression systems, yields are relatively low and the
system is expensive to run on a large scale; approximately 0.05 mg VLP per
1 liter cell culture can be routinely produced.
et al.
, 1993; Shanks & Lomonossoff, 2000; Singh
et al.
.. Plant-Based expression as a heterologous System
Production of proteins, especially pharmaceutically relevant proteins, in
plants is an important sector of biotechnology. Proteins can be expressed
in transgenic plants, in which the protein of interest is stably incorporated
into the plant genome. However, this is a time-consuming process and it can
take up to a year to generate transgenic plants that are stably expressing
the gene of interest. An alternative, and much faster and more flexible
system is to use plant viruses as expression vectors for high-level transient
expression (Awram
et al.
, 2002; Canizares
et al.
, 2005; Johnson
et al.
, 1997;
Porta & Lomonossoff, 1998; Scholthof
, 1996).
The advantages of heterologous protein expression in plants are high
yields, ease of purification, lack of mammalian contaminants (which is
important when producing pharmaceutically relevant proteins), ease of
manipulation, eukaryotic modification machinery, ease of scale-up, and
cost-effectiveness. The limitations of viral vectors are the high mutation
rates, which can lead to reversion to wild-type genomes and loss of the
foreign gene of interest. Also there are limitations to the size of the foreign
gene insert that can be tolerated by the virus.
There are several examples where VLPs have been produced using plant-
based expression systems. BMV VLPs have been produced using a TMV-
based expression vector (Choi & Rao, 2000). Also, CPMV- and PVX-based
expression vectors have been exploited to produce HBV
et al.
core particles in
plants (Mechtcheriakova
et al.
, 2006).
. ProduCtIon oF reCoMBInAnt Ads and AAV
The previous sections dealt with the production of bacteriophages, insect
viruses, and plant viruses. A few mammalian viruses have been exploited
for nanotechnology, such as recombinant Ad and AAV vectors, as well as
CPV,
virus. To date, the only systems extensively
used for biomedical nanotechnology are recombinant Ads and AAV. We will
outline routes of production of recombinant Ads and AAV in this section;
the reader is referred to the specialized literature regarding the production
of other systems (Fischlechner
Rubella
virus,
and
Influenza
et al.
, 2005; Singh
et al.
, 2006; Toellner
et
al.
, 2006).
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