Biomedical Engineering Reference
In-Depth Information
generating transgenic plants representing species
that were, at the time, intractable to other trans-
formation procedures. In the first such report, trans-
genic soybean plants were produced from meristem
tissue isolated from immature seeds (McCabe
et al.
1988). In this experiment, the screenable marker
gene
gusA
was introduced by particle bombardment
and transgenic plants were recovered in the absence
of selection by screening for
explant. However, the nature of the transformation
target is probably the most important single variable
in the success of gene transfer. The pretreatment of
explants with an osmoticum has often been shown
to improve transformation efficiency, probably by
preventing the deflection of particles by films or
droplets of water. Factors influencing the success of
gene transfer by particle bombardment have been
extensively reviewed (Sanford
et al.
1993, Birch &
Bower 1994).
-glucuronidase (GUS)
activity (Box 13.1). Other early successes included
cotton, papaya, maize and tobacco (Finer & McMullen
1990, Fitch
et al.
1990, Fromm
et al.
1990, Gordon-
Kamm
et al.
1990, Tomes
et al.
1990).
There appears to be no intrinsic limitation to
the scope of this procedure, since DNA delivery is
governed entirely by physical parameters. Many
different types of plant material have been used
as transformation targets, including callus, cell-
suspension cultures and organized tissues, such as
immature embryos, meristems and leaves. The
number of species in which transgenic plants can be
produced using variants of particle bombardment
has therefore increased dramatically over the last
10 years. Notable successes include almost all of the
commercially important cereals, i.e. rice (Christou
et al.
1991), wheat (Vasil
et al.
1992), oat (Somers
et al.
1992, Torbert
et al.
1995), sugar cane (Bower
& Birch 1992) and barley (Wan & Lemaux 1994,
Hagio
et al.
1995).
The original gunpowder-driven device has been
improved and modified, resulting in greater control
over particle velocity and hence greater repro-
ducibility of transformation conditions. An appara-
tus based on electric discharge (McCabe & Christou
1993) has been particularly useful for the develop-
ment of variety-independent gene-transfer methods
for the more recalcitrant cereals and legumes. Several
instruments have been developed where particle
acceleration is controlled by pressurized gas. These
include a pneumatic apparatus (Iida
et al.
1990), a
'particle inflow gun' using flowing helium (Finer
et al.
1992, Takeuchi
et al.
1992) and a device
utilizing compressed helium (Sanford
et al.
1991).
Physical parameters, such as particle size and accel-
eration (which affect the depth of penetration and
the amount of tissue damage), as well as the amount
and conformation of the DNA used to coat the par-
ticles, must be optimized for each species and type of
β
Other direct DNA-transfer methods
There is a great diversity of approaches for gene
transfer to animal cells and many of the same meth-
ods have been attempted in plants. Electroporation
has been used to transform not only protoplasts (see
above) but also walled plant cells, either growing in
suspension or as part of intact tissues. In many cases,
the target cells have been wounded or pretreated
with enzymes in order to facilitate gene transfer (e.g.
D'Halluin
et al.
1992, Laursen
et al.
1994). It has
been shown, however, that immature rice, wheat
and maize embryos can be transformed using elec-
troporation without any form of pretreatment (Kloti
et al.
1993, Xu & Li 1994). Other transformation
methods also involve perforation of the cell, including
the use of silicon carbide whiskers (Thompson
et al.
1995, Nagatani
et al.
1997), ultrasound (Zhang
et al.
1991) or a finely focused laser beam (Hoffman
1996). In most of these cases, only transient expres-
sion of the introduced DNA has been achieved,
although transgenic maize plants have been recov-
ered following whisker-mediated transformation.
Finally, microinjection has been used to introduce
DNA directly into the fertilized eggs of many animals
(Chapter 10). In plants, microinjection of DNA into
zygotes may also be the most direct way to produce
transgenics, but so far the technique is inefficient and
not widely used (Leduc
et al.
1996, Holm
et al.
2000).
In planta
transformation
Until recently, gene transfer to plants involved the
use of cells or explants as transformation targets and
an obligatory tissue-culture step was needed for the
regeneration of whole fertile plants. Experiments
using the model dicot
Arabidopsis thaliana
have led
