Date of publication: September 28,
2006
Source:
http://gmoinfo.jrc.it/gmp_browse_geninf.asp
Notification number:
B/ES/06/43
Member State:
Spain
Date of Acknowledgement
21/09/2006 00:00:00
Title of the Project
Field trial with transgenic Carrizo
citrange overexpressing a GA20-oxidase gene in sense and
antisense with the aim of modifying plant architecture,
flowering and fruiting behaviour, and to investigate the
performance of a non-transgenic variety grafted onto transgenic
lines.
Proposed period of release:
30/03/2007 00:00:00 to 30/03/2017
00:00:00
Name of the Institute(s) or
Company(ies)
Instituto Valenciano de Investigaciones
Agrarias., Generalitat Valenciana.;
3. Is the same GMPt release planned
elsewhere in the Community?
No
Has the same GMPt been notified
elsewhere by the same notifier?
No
Genetically modified plant
Complete name of the recipient or
parental plant(s)
|
Common Name |
Family Name |
Genus |
Species |
Subspecies |
Cultivar/breeding line |
| carrizo citrange |
rutaceae |
citroncirus |
citrus sinensis x
poncirus trifoliata |
|
|
2. Description of the traits and
characteristics which have been introduced or modified,
including marker genes and previous modifications:
The endogenous content of the
gibberellin GA1 has been altered in the transgenic plants by
overexpression of a GA 20-oxidase gene from citrus in sense or
antisense. As a consequence of the genetic modification, plant
architecture has been modified in the Carrizo citrange
transgenic plants.
All sense, antisense and transformed control plants are also
carrying a nptII transgene conferring resistance to kanamycin.
Genetic modification
3. Type of genetic modification:
Insertion;
In case of insertion of genetic
material, give the source and intended function of each
constituent fragment of the region to be inserted:
Cassette NOSpro: nptII:: NOSter: 1758
nucleotides. The nptII gene comes from Tn5 transposon of E.
coli. NOS regulatory regions (promoter and terminator) come from
the nopaline synthase (NOS) gene from Agrobacterium tumefaciens.
Transgene expression confers resistance to kanamycin to the
transgenic cells and tissues. This cassette is present in the
T-DNA from pBin19, which was used as the binary vector for A.
tumefaciens mediated transformation of citrange explants
(GenBank accesion of pBin19: nº U09365).
Cassette 35Spro 2X::CcGA 20-oxi1 sense or antisense:: 35Ster:
747, 1200 y 735 nucleotides, respectively. Promoter and
terminator regions come from the 35S gene from the cauliflower
mosaic virus (CaMV) and confer high and constitutive expression
of the transgene. The CcGA 20-oxi1 transgene comes from Carrizo
citrange, more specifically from the Poncirus trifoliata parent.
Its overexpression in sense or antisense induces an increase or
decrease, respectively, of the accumulation of GA1 in growing
shoots of the transgenic plants. Consequently, sense plants show
an elongated growth pattern compared to control plants.
Conversely, antisense plants are semi-dwarf and show shorter
flushes than control transformed with the empty vector (pBin19).
6. Brief description of the method
used for the genetic modification:
Co-cultivation of epicotyl segments
from Carrizo citrange with the disarmed strain EHA 105 of
Agrobacterium tumefaciens, and regeneration of whole plants from
the transgenic cells through organogenesis using proper tissue
culture media and conditions.
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:
Field trial of transgenic Carrizo
citrange overexpressing in sense or antisense a gene encoding a
GA 20-oxidase to study:
- modulation of tree architecture in plants growing in field
conditions. For this purpose, we propose to transfer to the
field 4 antisense lines (5A-8A, 5A-4, 5A 10 and 5A-33) and 3
sense lines (8S-8A, 8S-22 and 8S-23), plus one control line
(CT-81A; empty vector) (1 sample tree per line).
- flowering and fruit set behaviour in the same trees.
- potential growth modulator effect of the transgenic trees used
as rootstocks on a non-transgenic variety grafted onto them. For
this purpose, we propose to transfer to the field between 8 and
13 cuttings from lines: 5A-8A, 5A-4, 5A 10 and 5A-33, 8S-8A,
8S-22 and 8S-23, CT-81A and Carrizo citrange non-transgenic
control. All of them will be grafted with buds from
non-transformed Clemenules clementine. We will investigate
possible modulation of tree size, phenological characteristics
of the transgenic trees and fruit production and quality.
2. Geographical location of the site:
The release site will be located at the
experimental fields of the Instituto Valenciano de
Investigaciones Agrarias, Moncada, Valencia, Spain.
3. Size of the site (m2):
About 1000 m2.
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:
Notification B/ES/96/15. The experiment
has an extension of 1.638 m2. It has a total of 130 trees,
including 16 transgenic plants of Pineapple sweet orange, 16
transgenic plants of Mexican lime and 16 transgenic plants of
Carrizo citrange. In addition it has a similar number of
non-transgenic plants of the same species and an external border
of non-transgenic trees of Nules clementine. It has been
introduced in the plants´ genome the marker genes nptII and uidA
(GUS), that encode respectively for the neomycin
phosphotransferase II and ß-glucuronidase enzymes.
We are investigating: Morphological and phenological
characteristics of the transgenic trees, fruit quality,
expression of the transgenes in leaves, flowers and fruits,
stability of integration and expression of the transgenes,
transmission of the transgenes to the progeny, and possibility
of transgene dispersal through the pollen to non-transgenic
monoembryonic citrus trees surrounding the transgenic area.
Since 2001, the number of transgenic seeds expressing the uidA
gene has been yearly assessed in fruit produced by the border
trees.
As border trees are very close to the transgenic trees (3 to 6
m), our results indicate that possibilities of transgene
dispersal are very low in our conditions.
Environmental Impact and Risk
Management
Summary of the potential environmental
impact from the release of the GMPts:
Commercial citrus varieties are
propagated vegetatively by grafting of well-known genotypes onto
well-known rootstocks. In our Mediterranean conditions and
considering the citrus genotypes used, it is not possible that
transgenic plants could become weeds.
Citrus cultivars grown in the area are sexually compatible with
the transgenic plants. There are not wild citrus species and
relatives in Europe, so there are not possibilities of
compatible interactions between transgenic and wild plants.
Under natural conditions, cross-pollination between transgenic
lines and cultivated genotypes (citrus species or hybrids) is
theoretically possible. Pollination in citrus is exclusively
performed by insects, being bees the most successful
pollinators. However, there are laws in Valencia region
forbidding beehives where clementine is being planted. Moreover,
it is permitted to treat with insecticides against bees during
the citrus flowering season. This rule was adopted to avoid
cross-pollination and consequently seed production in fruits of
certain varieties. Presence of seeds in the fruit drastically
reduces its price, preventing possible commercialisation. Most
citrus species are parthenocarpic, meaning that they usually
produce fruit without seeds. At the IVIA area, clementine
orchards are very common, so there are not beehives in the
surroundings (neither at IVIA experimental fields), preventing
the possibilities of uncontrolled cross-pollination. In any
case, if cross-pollination occurs, transgenes will be only
expressed in the seed, which is never consumed.
Citrus varieties are not reproduced by seeds. In the incidental
case that transgenic seedlings could germinate in an orchard,
they would be removed by farmers as it is usually done with any
citrus seedling germinating in any orchard. Moreover, these
seedlings would never flower before being removed because citrus
seedlings need several years to start flowering.
Brief description of any measures
taken for the management of risks:
The field is being prepared according
to normal practices for citrus cultivation. Former trees were
removed and all their rests (including roots) were eliminated,
the soil has been treated and cleaned to avoid weeds and
phytopathological problems, it has been laser-levelled, and a
dropping watering system has been already installed. The
standard system of citrus cultivation will be used, removing
weeds and treating against pests.
We propose to plant in the field 4 antisense lines (5A-8A, 5A-4,
5A-10 and 5A-33) and 3 sense lines (8S-8A, 8S-22 and 8S-23),
plus a control line (CT-81A; empty vector) (1 sample tree per
line).
For the third objective of the trial, we propose to transfer to
the field between 8 and 13 cuttings from lines: 5A-8A, 5A-4,
5A-10 and 5A-33, 8S-8A, 8S-22 and 8S-23, CT-81A and Carrizo
citrange non-transgenic control. All of them will be grafted
with buds from non-transformed Clemenules clementine.
There will be about 90 trees in the field.
In most trees only the rootstock will be transgenic, and the 6
transgenic Carrizo citrange non-grafted trees will be planted in
the center of the field. In this way, we will favour that in
case of presence of bees during the flowering season transgenic
pollen will most likely be used to pollinate clementine flowers
from the 80 surrounding trees of the experimental field.
All rests coming from the transgenic trees of the field (whole
plants, rests from pruning, fruit, etc.) will be burned at the
IVIA incinerator.
Summary of foreseen field trial
studies focused to gain new data on environmental and human
health impact from the release:
Pollen dispersal frequency is being
assessed annually since 2001 in a transgenic field trial
authorized in 1997 (notification nº B/ES/96/15).
No experiment is being designed to study possible impact of the
transgenic fruit in human health.
Final report
-
European Commission administrative
information
Consent given by the Competent
Authority:
Not known |
