AgBioView - http://www.agbioworld.org, http://agbioview.listbot.com
Date: Jul 07 2000 06:06:34 EDT
From: Malcolm Livingstone
Thanks for your reply. Yes I agree that multiple copies are integrated and
multiple rearrangemnets are common. However my point is that as far as
field release or market release is concerned all these problematic lines
are eliminated. When I characterise my plants the only ones I keep are
single copy transformants with a Southern profile that gives confidence of
gene integrity. This is not foolproof but it doesn't have to be. The
cre/lox system can be used the way you suggest but retransformation is a
valid alternative. My comments were not meant to be comprehensive but just
an indication of workable approaches.
At 18:30 3/07/00 -0700, you wrote:
>A couple of minor corrections to your email.
>> How can we get rid of these genes after selection is complete and we
>> our transformed plants growing nicely in the glasshouse? There are a
>> of strategies that have been used successfully. Most are based on the
>> that it is possible to cross the transformed plants with untransformed
>> transferring the gene of interest (eg pest resistant gene) but leaving
>> selectable marker behind. This is because if genes are present in the
>> genome at different places they can be separated by traditional breeding
>> and selection. One approach is to use separate plasmids and directly
>> transfer them to the plant using microprojectile bombardment
>> (cotransformation). Because they are separate plasmids they will
>> randomly at different sites in the plant cell genome.
>In biolistic transformation most the separate plasmids integrate into a
>site and all the copies of however many plasmids (I've heard of up to
>end up tightly linked. The integration can be quit complex, with multiple
>copies of each plasmids including partial and rearranged copies. There is
>also some indication that host DNA also gets into this complex site. The
>details of how the integration occurs is very unknown. Our experience
>wheat is that only about 5% or fewer of the tranformed lines have multiple
>integration sites that can be segregated.
>> A more high-tech approach uses either the cre/lox, FLP/FRT or R-RS
>> (see Ow and Medberry, 1995). In this case one transforms the plant with
>> plasmid containing a sequence (lox) flanking the selectable marker. The
>> transformed plant is re-transformed with a gene (Cre) that recognises
>> excises all the DNA between the lox flanking sequences. The second
>> selectable marker and its cre gene are then removed using normal meiotic
>The Cre gene is usually in a separate transformed line that is then
>crossed to the line carrying the lox-containing construct. This allows
>excision of the marker followed by segregating away the Cre gene.
Subj: RE: Antibiotic marker genes
Date: Fri, 7 Jul 2000 12:41:49 PM Eastern Daylight Time
From: "Redenbaugh, Keith"
Greg is very correct about the food additive regulation for APH(3')II.
Calgene did extensive analysis on the marker gene and gene product. FDA
spent 4 1/2 years reviewing the data prior to its determination of food
additive status for APH(3')II (which of course is just another name for
NPTII). During this time period, FDA repeatedly came back to Calgene with
additional questions and data requirements. FDA also conducted a 3-day,
public Food Advisory Committee meeting in April 1994 to bring in outside
experts to examine any and all issues (food, feed, environmental) relating
FDA invited public comments from May 1 to July 30, 1991 and received only
47 responses, of which just 5 were negative. A second public comment
period from July 16 to August 16, 1993 resulted in no responses from the
public. The third public comment period was the Food Advisory Committee
the 15 comments, only 5 were negative.
Some FDA conclusions:
"FDA has concluded that the use of aminoglycoside
3'-aminophosphotransferase II is safe for use as a processing aid in the
development of new varieties ... intended for food use.
"No limits other than good manufacturing practice are needed to ensure the
safety of the petitioned use of APH(3')II."
There were two major results of FDA's review:
1) a determination that the kanamycin resistance marker gene and APH(3')II
were safe, and
2) a rigorous food additive-type model of safety assessment resulted,
which all companies have since followed.
Following are some references:
Redenbaugh, K., E. Emlay, W. Hiatt, C. Houck, J. Kiser, V. Knauf, M.
Kramer, J. Lindemann, L. Malyj, B. Martineau, K. McBride, D. Mitten.
Aminoglycoside 3'-phosphotransferase II (APH(3')II): Safety and Use in
Production of Genetically Engineered Plants. U.S. Food & Drug
Administration. Approved as food additive, May 23, 1994 (Fed. Reg.
Redenbaugh, K., W. Hiatt, B. Martineau, J. Lindemann, and D. Emlay.
1994. Aminoglycoside 3'-phosphotransferase II (APH(3')II): Review of its
safety and use in the production of genetically engineered plants. Food
Redenbaugh, K., T. Berner, D. Emlay, B. Frankos, W. Hiatt, C. Houck,
M. Kramer, L. Malyj, B. Martineau, N. Rachman, L. Rudenko, R. Sanders, R.
Sheehy, and R. Wixtrom. 1993. Regulatory issues for commercialization of
tomatoes with an antisense polygalacturonase gene. In Vitro Cell. Dev.
Biol. Plants 29P:17-26.
Redenbaugh, K. and J. Lindemann. 1994. "Safety determination of
the kanr marker gene for use in tomato, cotton and oilseed rape." In
Biosafety of Foods Derived by Modern Biotechnology. Proceedings Basel
on Biosafety 18 October 1994. BATS, Switzerland. 23-34.
Redenbaugh, K., W. Hiatt, B. Martineau, J. Lindemann, and D. Emlay.
1993. Aminoglycoside 3'-phosphotransferase II (APH(3')II): Safety and
in the production of genetically engineered plants. In WHO Workshop on
Health Aspects of the Use of Marker Genes in Plants and Possibilities for
their Use in Identification and Control of Genetically Modified Plants.
September 21-24, 1993. Rungsted, Denmark.
It is absolutely incredible that opponents of biotechnology claim that
biotech foods are untested and that they reject or ignore the complete and
rigorous safety DETERMINATION done by FDA on APH(3')II.
Date: Jul 06 2000 14:19:48 EDT
From: "James Reich"
Subject: Re: Stunting green progress
Financial Times, July 6 2000
STUNTING GREEN PROGRESS
A world environment body would create unecessary bureaucracy and divert
attention from existing goals, says Calestous Juma
At a recent meeting of the World Bank in Paris, Lionel Jospin, the French
prime minister, called on the United Nations to establish a world
environment organisation to create and enforce environmental regulations.
The call reflected the sense of urgency in addressing global environmental
problems. But such a proposal might delay, rather than hasten, action.
Advocates support the establishment of an agency for a number of reasons.
First, they attribute the UN's lagging environmental efforts to the fact
that environmental tasks are fragmented and performed by too many
unco-ordinated agencies and treaties. Second, they decry the lack of
enforcement mechanisms in most existing treaties - they claim that a new
agency would have more authority and would do a better job. Third they say
the agency would help transfer environmental technologies and finances to
developing nations. And finally, they appeal to the need for a
to the World Trade Organisation, which they consider inimical to
None of these reasons has a sound basis. The claim that consolidating
existing international organisations will result in a stronger and more
effective organisation, runs counter to the experiences of modern
institutions, which are decentralising their operations. Many UN agencies
are seeking to work through networks and a variety of institutional
alliances focusing on specific problems. Environmental problems are
diverse in character and require more specialised institutional responses.
Methods for combating global warming, for example, cannot be easily
applied to conservation of endangered species.
Environmental problems also cut across different sectors, and a world
environmental agency would need to cover every conceivable human activity.
It would be too cumbersome to work.
There is an urgent need to share experiences and lessons, codify
principles, promote guidelines and set new standards. But this is just
what global treaties on climate change, biological diversity, endangered
species and control of desert growth were created to do. The strength of
the treaties lies in the fact that they give more power and authority to
governments and citizens, not to centralised UN agencies.
It is true that most environmental agreements lack effective enforcement
mechanisms. One of the reasons is that governments cannot agree on how
they should work. Drawing on their experiences with the World Bank and the
International Monetary Fund, many developing countries are concerned that
a new environmental agency would only become another source of conditions
The real task is deciding how to get national governments to comply fully
with environmental laws. If governments promote greater compliance with
domestic environmental laws, they will find it easier to reflect this in
international agreements. What is perceived as deficient global
environmental regulations is really an indication of poor domestic
Most developing nations cannot meet their obligations under various
environmental treaties. They say this is partly because the richer nations
have not honoured their commitments to assist them with technology and
finances. There is no guarantee that the new agency will perform better in
In addition, developing nations have consistently argued that
environmental conservation should be promoted as part of their overall
economic goals, as agreed at the 1992 Earth Summit in Rio de Janeiro.
Creating a new agency focusing on environment over development, as is
proposed, would amount to
reneging on this historic agreement.
The claim that a new agency would serve as a counterweight to the WTO is
equally flawed. The WTO undoubtedly needs to take environmental issues
seriously. But this would be more effectively done by integrating
environment into trade activities, not simply by creating a new agency.
Unfortunately, the debate on creating a new agency diverts attention from
more urgent tasks. All nations need to do more to meet their obligations
under international treaties. Much of this involves domestic efforts to
cut pollution, protect wildlife, and conserve soils and freshwater.
Industrialised nations should also meet their international commitments by
sharing experiences and forging technology partnerships with developing
Tackling the world's environmental problems calls for urgent action. There
is no time to waste on launching a costly, politically divisive and
bureaucratic structure with no clear organising principle.
Mr Jospin's call is tantamount to saying a bigger Titanic will guarantee
the safe crossing of the Atlantic. It will not.
The writer is director of the Science, Technology and Innovation Programme
at the Centre for International Development at Harvard University