Biomedical Engineering Reference
In-Depth Information
Agitation in
liquid media
Low auxin +
low cytokinin
Shoots
Low auxin
+ high
cytokinin
High auxin
+ low
cytokinin
High
auxin
High
cytokin
Protoplasts
Cells
Callus
Plantlets
Solid
medium
Solid
medium
Roots
Maintenance
on solid
media with
low auxin
and low
cytokinin
content
Agitation in
liquid media
Enzymic digestion
shoots/roots
Callus
Cells
Protoplasts
Explant
on solid
medium
Agitation
in liquid
media
Fig. 12.4
Summary of the different
cultural manipulations possible with
plant cells, tissues and organs.
Enzymic digestion
nopaline, which are unusual amino acid derivatives
not found in normal plant tissue (Fig. 12.7).
The metabolism of opines is a central feature of
crown-gall disease. Opine synthesis is a property
conferred upon the plant cell when it is colonized by
A
.
tumefaciens
. The type of opine produced is deter-
mined not by the host plant but by the bacterial
strain. In general, the bacterium induces the syn-
thesis of an opine that it can catabolize and use as
its sole carbon and nitrogen source. Thus, bacteria
that utilize octopine induce tumours that synthesize
octopine, and those that utilize nopaline induce
tumours that synthesize nopaline (Bomhoff
et al.
1976, Montaya
et al.
1977).
Agrobacterium
-mediated
transformation
Crown-gall disease
Crown-gall is a plant tumour that can be induced in
a wide variety of gymnosperms and dicotyledonous
angiosperms (dicots) by inoculation of wound sites
with the Gram-negative soil bacterium
A. tumefa-
ciens
(Fig. 12.5). The involvement of bacteria in
this disease was established by Smith and Town-
send (1907). It was subsequently shown that the
crown-gall tissue represents true oncogenic trans-
formation, since the undifferentiated callus can
be cultivated
in vitro
even if the bacteria are killed
with antibiotics, and retains its tumorous properties
(Fig. 12.6). These properties include the ability to
form a tumour when grafted on to a healthy plant,
the capacity for unlimited growth as a callus in
tissue culture even in the absence of phytohormones
necessary for the
in vitro
growth of normal cells,
and the synthesis of opines, such as octopine and
Tumour-inducing (Ti) plasmids
Since the continued presence of
Agrobacterium
is not
required to maintain plant cells in their transformed
state, it is clear that some 'tumour-inducing prin-
ciple' is transferred from the bacterium to the plant
at the wound site. Zaenen
et al.
(1974) first noted
