based on SBSTTA 10 Submission February 2005
Any biological containment system setting out to prevent gene
flow of transgenes via pol en and seed must be 100% reliable and effective.
V-GURTs is likely to only be as good as its weakest link. There are a number of known events which can interfere with
At present there are a number of molecular containment
reliable performance of any of the 4 components employed by
strategies that aim to restrict gene flow either via pol en, seed
V-GURTs. Further research is required to address the
or sprouting of vegetative organs (e.g. tubers). Such strategies
include male sterility, maternal inheritance, seed sterility, prevention of sprouting, apomixis and temporal and tissue
Gene Silencing and epigenetic changes to DNA
None of these containment systems claims to have the
Gene silencing and epigenetic changes to transgenes
capacity to prevent gene flow both for pollen and seed except
have been observed repeatedly in transgenic plants,
for V-GURTs, also known as Terminator Technology.
especial y under stress conditions (Broer 1996, Meza et
V-GURTs is a genetic use restriction technology that – in its
2001). RNA-mediated silencing and DNA methylation
design - uses complex inducible gene expression systems to
are considered to have evolved as part of a host defense
make plants produce sterile seeds under specific conditions,
mechanism, active against viruses and parasitic DNA
which are active against transgenes (e.g. Riddihough and Pennisi 2001).
As no field trials or green house trials, nor any details of a complete and ful y functional V-GURTs plant have been
Recently, Srivastava and Ow (2003) for example, found
reported in the peer-reviewed literature, there is no scientific
that the site specific recombination system Cre/lox did not
data available to analyse the performance of this technology
perform as expected and the authors suggest that the Cre
and its reliability as a whole. Consequently a performance and
gene underwent a genetic or epigenetic change.
risk analysis for V-GURTs can only be carried out in part,
Whilst gene silencing of the repressor gene would lead to
focussing on the different individual genetic constructs and
sterile seeds, the silencing of either the recombinase or
components or the different expression systems for which
the toxin gene would result in viable seeds irrespective of
whether the inducer was applied or not. The potential
The basic design of V-GURTs
as detailed in US patent
silencing of the late embryonic abundance (LEA)
5,723,765 by the USDA and Delta & Pine Land is composed
promoter, which drives the toxin gene, is regarded by
Daniel (2002) as a crucial drawback of the terminator design as put forward by the USDA and Delta & Pine
• a cel toxin expressed in the late embryonic stage that wil
result in sterile seed. The toxin gene is inactivated by a spacer (short sequence of DNA), which can be removed
Reversibility of gene silencing (e.g. Mittelsten Scheid et
by a recombinase enzyme. To this effect, the spacer is
. 1998) may further add to unpredictabilities, e.g. seed
framed by recombinase recognition sites (e.g. lox sites for
• a recombinase enzyme that can activate the toxin gene, • a repressor protein that blocks the recombinase gene
Mutations frequently occur. Mutations of, for example, the
Once the inducer (e.g. tetracycline) has been applied, it wil
introduced recognition sites (e.g. lox sites) would result in
remove the repressor protein from the promoter of the
a permanently inactive toxin gene, leading to permanently
recombinase gene, thus recombinase is produced, which in
viable seeds, i.e. in this and al subsequent generations.
turn wil remove the spacer from the toxin gene. This now
Equal y, mutations to the recombinase promoter or gene
al ows the expression of the toxin in the late embryonic stage
would result in permanently viable seeds. Gene flow of
of the seed, kil ing the cel s and thus the growing seed.
There are other variants that use activators rather than
Loss of promoter activity:
repressors, but fol ow the same general principles outlined above.
Loss or reduction of promoter activity over time has been observed in a number of genetical y engineered systems.
The main components of V-GURTs are thus a) an inactive
The observed loss of promoter activity in the tetracycline-
gene coding for a cel toxin, b) an inducible site specific
inactivatable tTA expression system for example is
recombination system, c) an inducible expression system
presumed to be due to gene silencing (Tang et al.
using external inducers and d) an external inducer.
Leaking promoter systems:
disadvantage selective pressure wil favour genetic or epigenetic changes that lead to viable seeds or gene flow via
Many of the promoters in the inducible expression
pol en and capacity for reproduction, especial y for example
systems tested show a low level basal activity rather than
for transgenic trees. Effectiveness of V-GURTs applications
zero basal activity. For example, leakiness of the
may thus decrease over time and generations.
tetracycline-inducible promoter system was reported by De Veylder et al.
A further drawback of V-GURTs is that farmers growing conventional or traditional crops of the same species as the V-
Insufficient induction of promoter systems by inducing
GURTs variety in neighbouring fields wil still find their crops
contaminated by cross pol ination. Whilst this is a problem for
If the inducing agent, e.g. tetracycline, does not reach the
marketing their crops as GM free, especial y if the GM crops in
target cel s in sufficient quantity, the system wil only be
question were pharma crops, it may severely impact on food
partial y activated, resulting in viable seeds or in pol en
security. Farmers who save their traditional or conventional
capable of giving rise to viable seeds in neighbouring
seeds for replanting may find a significant percentage do not
crops. As stated by Daniel (2002), “it wil be difficult to
germinate as a result of cross pol ination, which in turn may
ascertain whether al the seeds treated with the
tetracycline inducer have triggered the gene switch (i.e.
whether tetracycline has penetrated al the seeds.)”
Unspecific or unintended induction of promoter system:
Many inducible promoters can be induced by more than
Broer I (1996). Stress inactivation of foreign genes in
one agent or by plant endogenous chemical agents. The
transgenic plants. Field Crops Research
AlcR based ethanol inducible system for example can be
Daniel H (2002). Molecular strategies for gene containment in
inappropriately triggered by endogenously produced
transgenic crops. Nature Biotechnology
20:581-586 – see
ethanol (due to anoxia) - (in Padidam 2003).
also Research Errata, Nature Biotechnology
Segregation of the different genetic components during
De Veylder L, Beeckman T, van Montagu M and Inze D
(2000). Increased leakiness of the tetracycline-inducible Triple-Op
promoter in dividing cells renders it unsuitable
It is crucial that functional components of V-GURTs and
for high inducible levels of a dominant negative CDC2aAt
the introduced GM trait remain linked during reproduction.
gene. Journal of Experimental Botany
For example, if separation resulting from segregation
Meza TJ et al
. (2001). The frequency of silencing in
should occur between the GM trait gene and the V-GURT
Arabidopsis thaliana varies highly between progeny of
genes, the GM trait may be passed on through seed and
siblings and can be influenced by environmental factors.
pol en to crops and weeds. Equal y, if any of the genes
involved in the V-GURTs system were segregated from
Mittelsten Scheid O et al. (1998). Release of epigenic gene
the others, the system would no longer function as
silencing by trans-acting mutations in Arabidopsis.
required. To avoid segregation, V-GURTs requires that all
Proceedings of the National Academy of Sciences, USA
the introduced genes be placed in very close proximity on
the same chromosome (strand of DNA) to create a linkage that wil reduce the likelihood of segregation as
Padidam M (2003). Chemical y regulated gene expression in
much as possible. If segregation occurs, any resulting
plants. Current Opinion in Plant Biology
gene flow of the transgenic trait (e.g. gene for Bt-
Riddihough G and Pennisi E (2001). The Evolution of
endotoxin, pharmaceutical component or lignin reduction)
Epigenetics. Science Magazine
293(5532) Srivastava V
to related cultivated, wild or weed relatives wil occur and
and Ow DW (2003). Rare instances of Cre-mediated
may be difficult to pick up in time to prevent it spreading
deletion product maintained in transgenic wheat. Plant
Srivastava V and Ow DW (2003). Rare instances of Cre-
mediated deletion product maintained in transgenic wheat.
Plant Molecular Biology
Tang W; Luo XY; Samuels V (2004). Regulated gene
To date, no functional and complete V-GURTs application has
expression with promoters responding to inducers. Plant
been detailed in the scientific literature. The evaluation of the
capacity of V-GURTs as a gene containment system
presented here has thus relied on the evaluation of its envisaged components.
A system can be only as good as its weakest parts. At
present, none of the components tested for any of the possible V-GURTs systems are 100% reliable or effective. Given that
at this stage the individual components of V-GURTs offer less than 100% efficiency or reliability, the combination of these components in one organism wil amount to still less. For example, if each of the 4 components performs to 95%, in combination their performance could reduce efficiency or reliability to as little as 81%.
Equal y, future evolution of V-GURTs must be taken into account. Because V-GURTs confer an evolutionary
THE CHEMICAL SENSES Relevant reading: Kandel & Schwartz, chapter 34. S Korsching, ‘Olfactory maps and odor images’ (pp. Montmayeur, & H Matsunami, ‘Receptors for bitter and sweet taste’ (pp. 366-371); both in Current Opinion in Neurobiology 12 (4) (August 2002). 1. Each olfactory receptor molecule, and by extension each olfactory receptor,responds in varying degrees to many
Beschwerde-Tagebuch ♂ 38 Jahre, 2009 Fange an zu Joggen. Stelle geringe Leistungsfähigkeit fest (schaffe nicht mal ein Programm für Ungeübte). Ziehende, sich geschwollen anfühlende Beine, geringe Leis-tungsfähigkeit beim Joggen (nach kurzer Zeit Ermüdung und Schmerzen in den Beinen, kaum Konditionsverbesserung), ge-ringe Kraft beim Krafttraining (Gewichtsreduzierung der Hanteln um 2