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Transgenic Lines Proven Unstable
The insert in every commercially approved GM line has undergone
rearrangement. The cauliflower mosaic virus promoter plays a major role. This
should be the final nail in the coffin for GM crops, says
Dr. Mae-Wan Ho, who has, for years,
challenged scientific committees advising governments over this very issue.
There is plenty of evidence that transgenic lines are unstable, which is
why ISIS has long recommended that appropriate molecular methods must be used
to document the stability of the GM insert before any transgenic line is
released into the environment. The characterization of the insert must be
‘event-specific’, which not only gives the structure of the insert,
but also the host genome sequences flanking the insert, proving that the insert
remains stable in successive generations. This recommendation has been
incorporated into the current European Directive (2001/18 /EC) on deliberate
release of GMOs.
But to this day, pro-GM scientists advising the UK and other governments
have refused to acknowledge the evidence on transgenic instability, and worse.
In its latest reply to ISIS, the UK Advisory Committee on Releases to the
Environment (ACRE) has gone as far as to say that event-specific molecular
characterization is not necessary, thus going against the European Directive
(see ISIS’ final response
to ACRE: Let the people decide ).
ISIS has reiterated 5 experiments which should be done to address the
‘areas of uncertainty’, one of which calls for full event-specific
molecular characterization of all transgenic lines to establish uniformity and
genetic stability of the transgenic DNA insert(s), and "comparison with
the original data supplied by the biotech company to gain approval for field
trials or for commercial release."
I am pleased to report that some effort has recently been made to do
such experiments by French scientists from the Laboratory of Methods for
Detecting GMOs in Versaille, and the Laboratory of Biometry and Artificial
Intelligence, Domaine de Vilvert in Jouy-en-Josas. And they have presented
their results in a poster at a conference in June 2003 [1].
The scientists recognized that, as labeling laws and thresholds are
established for foods containing GMOs in Europe, Japan, Australia, New Zealand
and elsewhere, "reliable GMO identification and quantification methods are
needed to comply with the regulations." And "in order for these tests to be
specific, the sequence and detailed characterization of the GMO inserts and
their edges are required."
Five different commercially approved GMOs in Europe were analyzed: three
from Monsanto, one from Bayer and one from Syngenta. All inserts were
rearranged from their intended gene order. Moreover, all five inserts showed
further rearrangements from the original structure submitted by the
companies. In other words, either the companies were mistaken about the
original structure, or more likely, further rearrangements had occurred
after the crops had been commercially grown. The details are given in Box
1.
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Box 1
Scrambling and further scrambling of GM inserts
T25 maize LibertyLink (Bayer)
Modified for tolerance to herbicide glufosinate. Company data
showed insert includes a truncated ampicillin resistance bla gene in the
plasmid vector pUC18, a CaMV 35S promoter (hereafter referred to as P35S)
driving a synthetic pat gene (glufosinate tolerance) terminated by CaMV
35S terminator (hereafter referred to as T-35S). On analysis, the insert was
found to have undergone further rearrangement, so that a second, truncated and
rearranged P35S has been joined to the 5’ (left, or head) end of the
insert, while additional pUC18 sequences were found at the 3’ (right, or
tail) end.
Edges flanking the insert show homologies (similarities) with
Huck retrotransposons (a class of mobile genetic elements) in the maize
genome.
Mon 810 maize YieldGard (Monsanto)
Modified for resistance to lepidopteran insects (butterflies
& moths). Company data showed insert has a P35S driving a CrylAb
synthetic gene with terminator T-nos. Analysis revealed however, that T-nos and
part of the 3’ (tail) end of the CrylAb gene have been deleted.
T-nos has been detected elsewhere in the genome, indicating that it has moved
from its original position.
The 5’ (head) end of the insertion site shows homology to
the long terminal repeats (LTR) of the maize alpha Zein gene cluster, but no
homology to the maize genome was detected at the 3’ site, indicating that
there has been scrambling of the maize genome at the insertion site.
GTS 40-3-2 soybean (Monsanto)
Modified for tolerance to herbicide glyphosate (Roundup Ready).
Company data showed insert with P35S driving a composite gene containing the
N-terminal chloroplast transit peptide (CPT4) joined to modified epsps
gene with T-nos terminator.Analysis revealed that a 254bp piece of DNA
homologous to the epsps gene and 534bp of unknown DNA have been joined
to the 3’end of the insert.
It was not possible to identify the insertion site at all,
indicating substantial genome scrambling or deletion at the insertion site.
Bt 176 maize (Syngenta)
Modified for tolerance to herbicide glufosinate, male sterility
and insect resistance. The structures of two inserts, originating from two GM
constructs, were provided by the company. Only the simpler construct was
analyzed. Company data showed insert contains P35S driving the bar gene
(glufosinate tolerance) terminated by T35S, followed by the ampicillin
resistance (bla) gene plus bacterial promoter, and plasmid origin of
replication, ori. Analysis revealed several fragments, all containing
CaMV 35S promoter, one with P35S joined to T35S, a second with P35S joined to
an unknown sequence, and a third with P35S joined to the bar gene with
the T35S deleted.
There were at least three insertion sites.
GA 21 maize (Monsanto)
Modified for tolerance to herbicide glyphosate (Roundup Ready).
Company data indicated insert contains multiple copies of the cassette with the
rice actin gene promoter (P-ract) driving the composite gene containing the
N-terminal chloroplast transit peptide (CPT4) joined to modified epsps
gene and T-nos. There were three complete cassettes flanked by a cassette with
P-ract partially deleted at the 5’ end, and one cassette with 3’
deletion of epsps plus a lone P-ract at the 3’end. Analysis found
partial deletion of P-ract and deletion of T-nos in two different
cassettes.
The insertion site at the 3’end is flanked by sequences of
pol polyprotein gene belonging to a PREM2-retrotransposon. |
The results revealed that,
- All GMO inserts had rearranged from the structure provided by the
company.
- Many of the breakpoints for rearrangement involve the CaMV 35S
promoter, as can be predicted from its known recombination hotspot.
- Scrambling of the genome at the site of insertion occurred in at
least two out of five inserts.
- GMO inserts appear to show a preference for mobile genetic elements
(retrotransposons), with Long Terminal Repeats containing strong promoters,
which would result in "altered spatial and temporal expression patterns of
genes" nearby. In addition, it increases the chances that the inserts will move
with the retrotransposons, resulting in further genome scrambling and
horizontal gene transfer.
With considerable irony, whether intended or not is unclear, the authors
conclude: "Studying GMO’s structure is necessary to develop reliable
quantification and detection tests complying with the different regulations,
but it also leads [one] to ask fundamental questions about genome
fluidity. Many of the mechanisms involved in recombinant DNA integration
are similar to those underlying genome evolution. Therefore,
characterized GMO inserts are a very good model to study the molecular
system involved in DNA rearrangements in general."
- Collonier C, Berthier G, Boyer F, Duplan M-N, Fernandez S, Kebdani N,
Kobilinsky A, Romanuk M, Bertheau Y. Characterization of commercial GMO
inserts: a source of useful material to study genome fluidity. Poster courtesy
of Pr. Gilles-Eric Seralini, Président du Conseil Scientifique du
CRII-GEN, www.crii-gen.org
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