Date of publication: January 16,
2006
Source:
http://gmoinfo.jrc.it/gmp_browse_geninf.asp
Notification number:
B/DE/05/168
Member State: Germany
Date of Acknowledgement: 20/12/2005
Title of the Project:
On the biological security of genetically modified cereals:
Effects of transgenic plants on beneficial fungal microorganisms
Proposed period of release From:01/03/2006
To:30/09/2008
Name of the Institute(s) or Company(ies):
Justus-Liebig-University;
3. Is the same GMPt release planned elsewhere in the
Community?
No
4 - Has the same GMPt been notified elsewhere by the same
notifier?
No
Genetically
modified plant
1. Complete name of the
recipient or parental plant(s)
|
Common Name
|
Family Name
|
Genus |
Species
|
Subspecies
|
Cultivar/breeding line
|
| barley
|
poaceae |
hordeum |
hordeum vulgare |
vulgare |
spring
barley/Golden Promise, Baronesse |
2. Description of the traits and characteristics which have
been introduced or modified, including marker genes and previous
modifications:
Two lines of GMPts, pYW210-9-(4001-4360) and
pJH271-Beta-Glu-307, have been notified for release into the
environment.
pYW210-9-(4001-4360) was transformed with a DNA sequence
encoding an endochitinase (cThEn42(GC)) of the soilborne fungus
Trichoderma harzianum, which is able to hydrolyze chitin of cell
walls of Rhizoctonia solani AG-8. It was shown to reduce growth
of the phytopathogenic fungi Rhizoctonia solani AG-8 and
Rhizoctonia oryzae in vitro. The gene sequence of this fungal
endochitinase used for plant transformation has been
codon-optimized to a G+C content of 65,1% to allow its
expression in barley. The bar gene (phosphinothricin
acetyltransferase) of the soilborne bacterium Streptomyces
hygroscopius is a marker during transformation and mediates
resistance against the herbicide Bialaphos (glufosinate
ammonium). Either gene is under control of the Ubi-1 promoter of
the ubiquitin gene of maize and, therefore, each gene is
expressed constitutively in all plant parts.
pJH271-Beta-Glu-307 was transformed with a DNA sequence encoding
a (1,3-1,4)-ß-glucanase, which has been generated by intragenic
recombination of two (1,3-1,4)-ß-glucanases of the bacteria
Bacillus amyloliquefaciens and Bacillus macerans. The gene is
under control of the hor-3 promoter of the D-hordein gene
(hor3-1) of barley and, therefore, is expressed during seed
development. Here, the (1,3-1,4)-ß-glucanase hydrolyzes
(1,3-1,4)-ß-glucans and, consequently, mobilizes endosperm
starch and protein to support initial seedling growth. The gene
sequence of the microbial (1,3-1,4)-ß-glucanase used for plant
transformation has been codon-optimized to a G+C content of 63%
to allow its expression in barley. The GMPt expresses two marker
genes, (i) a synthetic green fluorescent protein (sGFP) of
Aequorea victoria and (ii) the bar gene (phosphinothricin
acetyltransferase) of the soilborne bacterium Streptomyces
hygroscopius. sGFP mediates green fluorescence to transformed
cells when excited at a certain wavelength of light and is under
control of the CaMV35S promoter of the cauliflower mosaic virus.
The bar gene is a marker during transformation and mediates
resistance against the herbicide Bialaphos (glufosinate
ammonium). The bar gene is under control of the Ubi-1 promoter
of the ubiquitin gene of maize. Due to used promoters, either
marker gene is constitutively expressed in all plant parts.
Genetic
modification
3. Type of genetic
modification:
Insertion;
4. In case of insertion of genetic material, give the source
and intended function of each constituent fragment of the region
to be inserted:
pYW210-9-(4001-4360)
The endochitinase (cThEn42(GC)) of the soilborne fungus
Trichoderma harzianum is known to hydrolyze fungal cell walls
and, therefore, is expected to enhance plant resistance against
fungal diseases.
The bar gene (phosphinothricin acetyltransferase) of the
soilborne bacterium Streptomyces hygroscopius is a selectable
marker during transformation and mediates resistance against the
herbicide Bialaphos (glufosinate ammonium).
Both genes are under control of the Ubi-1 promoter of the
ubiquitin gene of maize and, therefore, are expressed
constitutively in all plant parts.
pJH271-Beta-Glu-307
Hybrid (1,3-1,4)-ß-glucanases of the bacteria Bacillus
amyloliquefaciens and Bacillus macerans are under control of the
hor-3 promoter of the D-hordein gene (hor3-1) of barley and,
therefore, are expressed during seed development. The gene
mediates an enhanced hydrolization of (1,3-1,4)-ß-glucans and,
consequently, mobilization of endosperm starch and protein to
support initial seedling growth. In addition, glucans, but not
(1,3-1,4)-ß-glucans, are components of fungal cell walls and
might be hydrolyzed by the recombinant protein.
Synthetic green fluorescent protein (sGFP) of Aequorea victoria
mediates green fluorescence to transformed cells when excited at
a certain wavelength of light. The sGFP gene is under control of
the CaMV35S promoter of the cauliflower mosaic virus. Therefore,
the gene is constitutively expressed in all plant parts.
The bar gene (phosphinothricin acetyltransferase) of the
soilborne bacterium Streptomyces hygroscopius is used as marker
during transformation and mediates resistance against the
herbicide Bialaphos (glufosinate ammonium. The bar gene is under
control of the Ubi-1 promoter of the ubiquitin gene of maize.
Therefore, the gene is constitutively expressed in all plant
parts.
6. Brief description of the method used for the genetic
modification:
Immature embryos were transformed by cocultivation with
Agrobacterium tumefaciens strain AGL-1 harboring the constructs
pYW210 or pJH271. After cocultivation, immature embryos were
transferred to callus inducing medium containing the herbicide
Bialaphos. Subsequently, transformed calli were transferred to
shoot inducing- and root inducing media to obtain the
genetically transformed plants.
7. If the recipient or parental plant is a forest tree
species, describe ways and extent of dissemination and specific
factors affecting dissemination:
not applicable
Experimental
Release
1. Purpose of the release:
The purpose of the release is (i) to investigate the
influence of the recombinant enzymes on GMPts-Glomus
intraradices interactions, considering G. intraradices as an
agricultural relevant representative of mutualistic soilborne
fungi, and (ii) to perform epidemiological examinations
regarding the colonization of the GMPts by naturally occurring
parasitic fungi. The extent of mutualistic and parasitic
infestation on GMPt will be evaluated.
2. Geographical location of the site:
Field (Flur)/cadastral parcel (Flurstück) 15/75/2, Alter
Steinbacher Weg 44, 35394 Giessen, District (Landkreis) Giessen,
Hessen, Germany
3. Size of the site (m2):
12 m2 of genetically modified plants
4. Relevant data regarding previous releases carried out with
the same GM-plant, if any, specifically related to the potential
environmental and human health impacts from the release:
Both transgenic lines, pYW210-9-(4001-4360) and
pJH271-Beta-Glu-307, have been notified for release and have
been released in 2005 at various field stations at the
Washington State University, Pullman (USA)(Notification numbers
05-035-13n (Th42-2) for pYW210-9-(4001-4360)and 05-035-14n (di
04-029-02n) for pJH271-Beta-Glu-307)in order to propagate and to
backcross both transgenic lines with high yielding cultivars.
Indicated notification numbers have been given by the “Animal
and Plant Health Inspection Service – APHIS”, Riverdale
(Maryland), USA.
Due to the nature of these releases neither positive nor
negative effects of the transgenes on target or non-target
organisms have been recorded. GMPts and respective parent
cultivars were comparably colonized by aphids and their
beneficial antagonists, GMPts showed the same characteristics
regarding flowering, pollination and seed development and GMPts
did not differ in survivability and straw decomposition in the
field compared to their respective parent cultivars.
All inserted marker genes have been tested in feeding
experiments and did not show any harmful impact on fed
organisms. The substrates ((1,3-1,4)-ß-glucans and chitin) which
are hydrolyzed by the respective recombinant enzymes
((1,3-1,4)-ß-glucanase and endochitinase) do not exist in
mammalian organisms.
Albeit chitin is found in the cuticula of insects, it is not
present in the outermost layers of the integument and, in inner
layers, is tightly bound to proteins. Therefore, any chitinases
(transgenic or plant chitinases) have no or limited access to
chitin of insects. The portion of chitin present in peritrophic
matrices lining up the gut epithelium of insects is rather low
(3-13%) and not expected to be affected by the endochitinase
since especially herbivoric insects are not impaired by plant
chitinases. Consistently, the above described GMPts,
pYW210-9-(4001-4360) and pJH271-Beta-Glu-307, were similary
infested by aphids as compared to their respective parental
cultivars when grown in the field at the Washington State
University, Pullman (USA). The experiences in releasing both
transgenic lines made by scientifically educated staff of the
CAHNRS of the Washington State University, Pullman (USA), base
on qualitative observations.
All transgenes expressed in both transgenic lines do not show
any homology to known toxins and allergens.
All observations and experiences made, combined with the
knowledge on the recombinant enzymes and the distribution of the
respective substrates, do not indicate any direct or indirect
impact on the environment or human health from the release.
The impact of the GMPts on arbuscular mycorrhizal fungi as
mutualistic organisms with agricultural relevance can not be
excluded. For that reason, one main focus of the release is the
monitoring of GMPts-Glomus intraradices interactions.
Environmental
Impact and Risk Management
Summary of the potential
environmental impact from the release of the GMPts:
There is no scientific reason to assume that the traits
introduced into either transgenic line could directly or
indirectly confer increased advantages in natural environments
or that a significant environmental benefit is expected. None of
the traits introduced into either transgenic line has any impact
on traits that are related to distribution, pollination,
reproduction or survivability of barley in natural environments.
Due to domestication, the cultivation and distribution of barley
is entirely depending on anthropogenic agricultural practices.
Taken together, the potential environmental impact from the
described GMPts have to be regarded as similar to that from
conventional barley cultivars.
pYW210-9-(4001-4360)
The introduced endochitinase (cThEn42(GC)) is expected to
enhance plant resistance against fungal diseases because of its
ability to hydrolyze chitin of fungal cell walls. In case that
GMPts harboring the recombinant endochitinase are more resistant
to fungal pathogens, these plants are expected to gain an
increased selective advantage in agricultural environments when
confronted with a high disease pressure. Apart from fungi,
chitin is a component of inner layers of the cuticula of
insects. Other parts of insects which are potentially accessible
to transgenic or natural occurring chitinases are tightly bound
to proteins and, hence, are not expected to be hydrolyzed by
chitinases.
The bar gene (phosphinothricin acetyltransferase) mediates
resistance against the herbicide Bialaphos (glufosinate
ammonium) and is not expected to confer an increased selective
advantage in natural environments.
pJH271-Beta-Glu-307
The introduced (1,3-1,4)-β-glucanase is expressed during seed
development and mediates an enhanced hydrolyzation of
(1,3-1,4)-β-glucans in germinating seeds. It is not known
whether the recombinant enzyme is able to hydrolyze glucans of
fungal cell walls, thereby mediating resistance to fungal
pathogens. In case, pJH271-Beta-Glu-307 is more resistant to
fungal pathogens, these plants are expected to gain an increased
selective advantage in agricultural environments confronted with
a high disease-pressure. However, the introduced
(1,3-1,4)-β-glucanase is spatio-temporally expressed during seed
development and the enzyme possesses an high substrate
specificity to (1,3-1,4)-β-glucans of the seed endosperm and
aleurone.
The introduced synthetic green fluorescent protein (sGFP) of
Aequorea victoria mediates green fluorescence to transformed
cells when excited at a certain wavelength of light and does not
possess any physiological function in the GMPts.
The bar gene (phosphinothricin acetyltransferase) mediates
resistance against the herbicide Bialaphos (glufosinate
ammonium) and is not expected to confer an increased selective
advantage in natural environments.
Brief description of any measures taken for the management of
risks:
Barley is a self-pollinating plant with a limited ability to
cross-pollination. Cross-pollination is a rare event and
diminishes by distance between two plants. In order to prevent
any cross-pollination, transgenic field plots are framed by a
conventional barley variety (width: 5 m) that serves as a pollen
trap. This barley is surrounded by a band without plants (width:
5 m), which is finally framed by dicotyledonous plants (width:
25 m).
Procedures performed to limit seed and pollen dispersal:
• Potential pollen acceptors (wild barley, Elymus sp., cereals)
occurring within the site of the field trial will be eliminated
throughout the ongoing field experiments. To this end, the field
site will be regularly observed for respective plants.
• Plots of transgenic and non-transgenic plants will be
harvested by hand to reduce the dispersal of seeds. Harvested
seeds will be stored in certified S1 gene laboratories.
• Plants of the pollen trap barley culture will be harvested by
a combine harvester and harvested seeds will be destroyed.
• Plants growing within the site of field trials that are not
forwarded to analyses will be destroyed by application of a
non-selective herbicide. Dead plant material will be chopped and
covered with soil.
• The field will be observed for volunteer plants growing for at
least one year after finishing all experiments. In case
volunteer plants have been identified, the observation will be
automatically extended for another year.
Summary of foreseen field trial studies focused to gain new
data on environmental and human health impact from the release:
The field trials are designed to investigate the
mycorrhization of either transgenic line and the respective
parent cultivars by the arbuscular mycorrhizal fungus Glomus
intraradices. Therefore, one half of the plot sites will be
inoculated with the mutualistic G. intraradices prior to
planting transgenic and non-transgenic seeds. Mycorrhization as
an important agricultural aspect in crop production will be
monitored in randomly selected plant roots.
A second aspect will follow the occurrence of fungal diseases on
selected transgenic and non-transgenic plants. The aim is to
record whether transgenic or non-transgenic plants are
preferably infected by naturally occurring fungal pathogens.
Final report
-
European
Commission administrative information
Consent given by the Competent
Authority: Not Known |
