Technical Opinion no. 1596/2008

Proceedings:  01200.002293/2004-16
Applicant:  Monsanto do Brasil Ltda.
CNPJ:   64.858.525/0001-45
Address: Avenida Nações Unidas, 12901, Torre Norte, 7º e 8º Andares, 04578-000 São Paulo, SP.
Matter: Commercial Release of Genetically Modified Corn.
Previous extract: 134/2004. Published on 06.09.2004
Meeting: 116th Regular Meeting held on September 18, 2008.
Decision: GRANTED.

CTNBio, following approval  of an application for Technical  Opinion related to commercial release of genetically modified glyphosate tolerant corn Roundup Ready 2 Event NK603, as well as all progenies originated from the  NK603  transformation event  and its derivatives of crossing of non transgenic corn lines and populations with lines carriers of event NK603, was favorable to the GRANTING under the terms of  this technical opinion.
Monsanto do Brasil Ltda. requested a CTNBio Technical  Opinion for the free registration, use, essays, tests, sowing, transportation, storage, marketing, consumption, import, release  and discarding  of  glyphosate tolerant corn (Zea mays, L.) Roundup Ready 2 Event NK603. NK603 corn  expresses glyphosate protein CP4 5-Enol-pyruvylshikimate-3-phosphate synthase (CP4 EPSPS). Weed control performed by glyphosate is due  to  inhibition of  EPSPS enzyme naturally produced by the plant. The enzyme catalyzes  a  critical phase  of  the metabolic way of shikimic acid for biosynthesis of aromatic amino acids in plants  and microorganisms. CP4 EPSPS proteins have low glyphosate affinity, when compared with the wild EPSPS protein. Therefore, when the NK603 corn expressing protein CP4 EPSPS is treated with glyphosate, the plants keep developing normally. The continuous action of glyphosate tolerant CP4 EPSPS enzyme catalyzes the synthesis  of amino acids required for the normal development of the plant. The biosynthetic way of aromatic amino acids  is not  found in animals, which explains the glyphosate  selective activity in plants  and contributes towards low toxicity to mammals.  Two cp4 epsps  gene expression cassettes were introducing  in the corn genome  through a single insert, producing  NK603 corn. The cp4 epsps  gene  is derived from a bacterium common  in the soil, Agrobacterium sp. strain CP4, which codifies  the expression of  protein EPSPS naturally tolerant  to  glyphosate. The gene donor organism, A. tumefaciens strain CP4 is a  bacterium commonly found  in the soil and causes galls  in susceptible  plants, with no scientific  evidence indicating that  it may cause adverse effects to humans or animals. Toxicity tests  were conducted with EPSPS-synthase enzyme  isolated from transformed  plants. Ingestion through gastric gavage of the protein molecule in doses  1000 times above the figure for modified seeds failed to cause any physiologic change in animals tested. Results of in vitro proteolysis also verified rapid digestion and innocuousness of the engineered protein, removing any suspicions of allergenicity.  CP4 EPSPS protein is atoxic, as demonstrated by a oral acute toxicity  stud, when CP4 EPSPS was administered to mice in one single high dose. The protein, produced and purified from Escherichia coli, was characterized and showed to be equivalent  to the CP4 EPSPS produced in the corn. The purified  protein was orally administered to mice for assessing its acute toxicity. Acute administration was deemed as  appropriate in assessing  the safety of CP4 EPSPS, since toxic proteins act by means of acute mechanisms. The no-observed-effect-level (NOEL)  for oral toxicity in mice was 572  mg/kg, the highest tested dose. The result represented a safety margin of about 260,000 times,  based on the average daily consumption of corn in the United States and average expression of the protein in the glyphosate tolerant genetically modified corn (assuming that there is no  loss of CP4 EPSPS  throughout processing). There were no statistically different differences in body weight, aggregate body weight or food  consumption between control  groups (with the  vehicle or bovine albumin  serum) and groups treated with the purified CP4 EPSPS protein. EPSPS proteins are ubiquitous in nature and  are  naturally present in foods derived from plant and microbial sources and do not  show significant homology of  amino acids with proteins known to be toxic  or allergenic to mammals. Apparently, the CP4 EPSPS protein was rapidly digested by in vitro gastric and bowel fluids. Besides, EPSPS  proteins that  have a history of safe use  are widely used  in  the diet, corn is  a plant unable to  survive  in natural  conditions,  without  technical assistance. Therefore there is no possibility that the corn shall be transformed in an  invading plant or pest. Even in case of un unlikely genic escape, the possibility of fixation of an allele  containing  the genic sequence that  grants tolerance to glyphosinate  in the population is  very reduced  in the absence of selective pressure. The NK603 corn demonstrated to be equivalent to conventional corn, with the exception of its tolerance to glyphosate. Its basic interactions with other living organisms in the environment are not deemed to be different from the ones of conventional corn. Though there is a potential exposure of weed, pests and pathogens of corn culture to the  CP4 EPSPS and CP4 EPSPS L214P proteins that are expressed in the  NK603 corn, there are no concerns that the process may negatively affect such populations. Through trophic transfer and decomposition processes, organisms that  are not targets to CP4 EPSPS and CP4 EPSPS L324P proteins, such as predators and preys  of corn pests, may be exposed  to very low levels of such proteins without, however, emerging any evidence of negative effects on the non-target organisms. Environment safety of the EPSPS  protein  family is widely accepted, since the  proteins are  ubiquitous in nature (bacteria, fungi, algae and higher plants), do not  have known  toxicity, are not associated to pathogenicity and fail  to grant any selective advantage to plants that  contain the proteins. For the foregoing, there are no restrictions to the use of  the genetically modified corn and its  derivatives, either as human of animal food. In addition, the proteins stem  from the EPSPS protein family, with a long history of safe consumption and exposure, occurring ubiquitously in plants and microorganisms. Analytic studies comparing the composition of  kernel and forage of NK603 with  the conventional corn show that the  genetically modified glyphosate tolerant NK603  corn is substantially equivalent to  conventional corn. Based on studies conducted during the several years of marketing NK603 corn and other glyphosate tolerant cultures expressing the CP4  EPSPS protein, coupled with the safe  history of the  corn as a consumption source for human and animal food, one reaches  the  conclusion that glyphosate  tolerant NK603 corn, or Roundup Ready 2 corn  is substantially equivalent and as safe as conventional  corn. Coexistence of cultivars of conventional  (either improved or wild)  corn and  cultivars of transgenic corn is possible  from the agronomic viewpoint and shall comply with the provisions of CTNBio Ruling Directive no. 4. According to Article 1 of Law no. 11,460, of March 21, 2007 ”research and  cultivation of  genetically modified organisms may not be conducted in  Amerindian areas and conservation units”. Regarding the scope of Article 14 of Law no. 11,105/05, CTNBio holds that the request complies with applicable legislation and regulation aimed at securing biosafety of the environment, agriculture, human and animal health.
 CTNBio Technical Opinion
1. GMO Identification
GMO name:  Roundup Ready 2 Corn – Event NK603
Applicant:  Monsanto do Brasil Ltda.
Species:  Zea mays – Corn
Inserted  Feature: Glyphosate herbicide tolerance
Insertion method: Biobalistics  (particle acceleration) method
Prospective use: Production of silage and kernel for human and animal consumption  from the GMO and its derivatives
II. General  Information
Zea mays L., corn, is a species of the Maydae tribe, included in the subfamily Panicoidae, family Graminea (Poacea). Genera belonging to the Maydae tribe  include Zea and Tripsacum in the Western Hemisphere.  Corn is a separate species within the Zea subgenus, with a chromosome number 2n = 20,21,22,24(9). The wild  species closest to corn  is teosinte, found in Mexico and  some other places of Central America, where it may cross with cultivated corn in production fields. Cultivated corn may also cross with a more  distant genus, the  Tripsacum.  The crossing seldom happens and results in a sterile male progeny.
Corn has  a history of eight  thousand  years in the Americas. Out of all cultivated plants, corn is probably the  one possessing the largest genetic variability.  Today, about three hundred races of corn are identified and, within each such race, there are thousands of cultivars. Corn is currently the cultivated species that reached the highest degree of domestication and it may only survive in nature when raised by man(4). Normally, the maintenance of  this genetic variability has been achieved through individualized storage, in germplasm banks, under  controlled humidity and temperature. There are several germplasm banks,  in Brazil and all over the world. Embrapa, the Brazilian Agricultural Research Agency, has two germplasm banks,  one at Embrapa Recursos Genéticos e Biotecnologia, Embrapa Genetic Resources and Biotechnology, in Brasília, Federal District, Brazil, and another  at Embrapa Milho e Sorgo, Embrapa Maize and Sorghum, in Sete Lagoas, Brazil. Corn is farmed in over 100 countries, with a total estimated production of 705  million tons per year.
Corn is one of the most important food  sources in the world and  is the raw material  for a large range  of products. From 70% to 80% of corn produced in Brazil is consumed by the swine and poultry productive chain.
Brazil is the world third largest corn producer, with  an output of about 35 million tons  in 2005, behind the United States of America (282 million tons) and China (139 million tons)(11).In Brazil, corn is planted basically in two different crops (summer and safrinha, or  small  crop) and is cultivated in practically all the domestic territory, with 92% concentrated in the Southern Region (47% of production), Southeastern Region (21% of production) and Center Western Region (24% of production)(8).
Corn is one of the most efficient plants  in converting solar energy in food  and is used as raw material for several products. The increase  in corn consumption exceeded 100 million tons between 1993 and 2001, representing an average  yearly increase  of 11.1 million tons per  year. A large part of this increased production was due  to genetic  improvement, leading to  ears containing about  1,000 seed-corns. Increased corn production and consumption all  over the world is associated to its multiple uses, population growth,  changes in feeding habits, and growth in the number of farmed swine and  poultry.
Brazil is the world third largest consumer of pesticides. The country has currently 142  pesticides  registered for corn. There are several reports of resistance cases caused by the constant and indiscriminate use of herbicides in corn farming in Brazil. According to CUT(24) citing data from the  World  Health Organization (WHO), one million people are intoxicated each year by farm pesticides in Brazil. The Ministry of Health confirms that  in sixteen Brazilian states, farm defensives  are the largest cause affecting farmers’  health.
The use of herbicide tolerant corn, such as  NK603 corn, has given an opportunity for efficient management of pests and rotation of  herbicide principles in corn farming(13). One impact recorded is the reduction of herbicide use in farming pre-emergence. Farmers have  reduced the rates of pre-emergent herbicide applied to the soil  and, through the use of wide spectrum herbicides, as glyphosate, have managed to control pests  in an effective way. Replacement of several  herbicides  by a  single  herbicide with a wider spectrum has resulted in savings around  $25/ha. Studies conducted in the United States with herbicide tolerant  corn suggest positive economic yields  with the  single use of glyphosate or glyphosate associated to conventional herbicides in  pre-emergence(25). The results establish that herbicide tolerant corn, including NK603, provides an increase in productivity while reducing herbicide costs(7).
Glyphosate  tolerant NK603 corn was  released for the first time in the United States for farming and marketing in 2000,  and is currently farmed, or has its consumption permitted, in  twelve countries: Argentina, Australia, Canada, China, European Unit, Japan, Korea, Mexico, Philippines, South Africa, Taiwan and USA. NK603 corn expresses CP4 5-Enol-pyruvylshikimate-3-phosphate synthase (CP4 EPSPS) proteins that are glyphosate tolerant. CP4 EPSPS protein is one of several EPSPS proteins found in nature, which are produced by  plants, bacteria and fungi, but not by animals, since  those do not possess  the metabolic way for its synthesis. Therefore, different versions of the EPSPS protein  are normally present  in all food derived from plants and microorganisms. The gene donor  organism, Agrobacterium tumefaciens strain CP4, is a bacterium normally found in the  soil. It causes galls  in susceptible plants and there is no scientific evidence indicating that it may cause adverse  effects in humans or animals.
It  is worth noticing that the history of farming, marketing, use and experience with other genetically modified cultures that express CP4 EPSPS  protein since  the first marketing  of RR soy  in 1994,  has shown that  the protein failed to display  any risk to the environment, and to human and animal  health.
III. Description of the  GMO and Proteins  Expressed
Gene cp4 epsps, which codifies  a  glyphosate  tolerant form of  the 5-Enol-pyruvylshikimate-3-phosphate synthase (EPSPS) was isolated from  the bacterium Agrobacterium tumefaciens strain  CP4 and  inserted  in corn through biobalistics (particle acceleration). The  action of glyphosate, causing the death of plants, takes place  due to  its ability to block the activity of the target  enzyme (EPSPS) belonging  to the biosynthetic way of aromatic  amino  acids tyrosine, phenylalanine and tryptophan. Thus, plant cells that  express protein CP4 EPSPS keep producing aromatic amino  acids  essential to their metabolism, even in the  presence of glyphosate. Protein CP4 EPSPS is one of several EPSPS  proteins found  in nature, produced by plants, bacteria and fungi, but not by animals, which lack the metabolic way for the protein synthesis. Therefore, different versions of  the EPSPS protein are normally present in all food derived from plants and microorganisms. The gene donor organism, Agrobacterium tumefaciens strain CP4, is a bacterium normally found in the  soil that causes galls  in susceptible plants and there is no scientific evidence indicating that it may cause adverse  effects in humans or animals.
NK603 corn was produced through genetic transformation of a LH82XB73 corn lineage using a DNA fragment of 6706 pairs  of  bases (pb) containing  two adjoining cassettes of  gene cp4 epsps to  express the CP4  EPSPS protein.  Each such cassettes, namely, the proximal cassette (closer to the 5’ end) and distal cassette (closer to the 3’ end) contained a single copy of the cp4 epsps gene and regulating sequences.  In both cassettes, sequences of the cp4 epsps genes were linked to transit polypeptide sequences to chloroplast (CTP2) obtained from an epsps gene of Arabidopsis thaliana. The function of transit polypeptides is transporting the CP4 EPSPS protein to the chloroplasts where  the metabolic way responsible  for synthesizing aromatic amino acids operates. CTP4s are removed from protein CP4 EPSPS after its delivery at the chloroplast. In the proximal  cassette,  fragment ctp2-epsps was placed under the control of  rice promoter actin1  and its intron and, in the distal cassette, placed under the control  of CaMV 35S modified promoter (e35S). At the distal cassette, between promoter e35S and the CTP2 sequence, an intron  of  0.8 kb, from a corn protein, was also  introduced, involved in answers to thermal shocks (hsps70)  with the  purpose of  increasing the levels of genetic transcription. In both cassettes, sequences cp4 epsps were connected to the sequence of 0.3 kb  of nopaline synthase 3’ (not  translated)  named NOS 3’, with the purpose of providing a sign  for polyadenylation of the messenger RNA (mRNA). The DNA fragment of  6706 pb was used  in the genetic transformation was isolated  from plasmid PV-ZMGT32L as a single fragment through digestion with a restriction enzyme MIuI and separation in electrophoresis gel. Therefore, this fragment did not contain other plasmid elements as origin of plasmid replication and the sequence of gene npt II codifying the neomycin  transferase type II  enzyme. This enzyme grants resistance  to aminoglycoside antibiotics such as  kanamicyn and neomycin used to select the bacteria during the  construct and  multiplication  of the plasmid. Transformed cells were  selected in tissue culture in the presence of  glyphosate.
The DNA fragment was characterized using analyses of Southern Blot, a Polymerase Chain Reaction (PCR) technique, and sequencing of the inserted fragment and its bordering regions in the transformed corn genome. Results showed  that the NK603 corn genome  contains a single  exogenous DNA insertion and that  no  DNA of the plasmid replication structure had been detected in the NK603 corn genome. Inside  the single  insert, a complete copy of the  6706 pb  DNA fragment used in the transformation was found and  one  217 pb fragment  in the region of  the actin promoter that fails to  contain the necessary elements to act as a promoter. Both the proximal and distal cassettes of  the cp4 epsps gene are present in the single insert and their genetic  components are intact.  In the distal cassette, the  cp4 epsps  gene nucleotide sequence differs from the original sequence used  in  the process of transformation in two nucleotides. One nucleotide change was silent while the  other resulted in the substitution of an amino acid in position 214. The changed nucleotide in position 214 pb resulted in the coding of a leucine instead of a proline. This new sequence came to be called  cp4 epsps L214p. Analyses of PCR  products of terminal 3’ of the   inserted DNA revealed  the co-integration of  a 305 pb  additional DNA fragment of a chloroplast DNA. Results of computational biology showed that  the co-integrated  DNA corresponds to a part of DNA sequences  codifying the  alpha subunit of RNA polymerase and the ribosome protein S11. It is believed that  the origin of  this DNA is the chloroplast of the transformed cell.
Proteins CP4  EPSPS and CP4 EPSPS L214P are present in small concentrations in  seeds and forage of NK603 corn since rice actin promoters and e35s act in a constitutive  way. Studies show that there is equivalence of both  with proteins  produced  in Escherichia coli containing plasmid of  heterologous  expression possessing homology with EPSPS proteins that are naturally produced by plants and microorganisms used in human and  animal food. Regarding nutritional and environmental safety, both the EPSPS proteins naturally present in non-transgenic plants and microorganisms and the CP4 EPSPS proteins expressed  in glyphosate tolerant genetically modified  cultures  belong to a family of proteins known for their absence of toxicity, non association to pathogenicity  events and failure to grant any selective advantage  to  plants or microorganisms containing them. Although primary amino acid sequences  in  different members  of the EPSPS protein family show considerable divergence,  the expressed proteins are highly related in terms of  structure and function.
IV. Aspects related to  Human and Animal Health
Security assessment of food derived  from genetically modified  raw-materials is based on risk analysis, a scientific methodology that encompasses  the phases of risk assessment, risk management and risk communication. In the risk assessment, one pursues the qualitative and quantitative characterization of potential adverse effects,  based  on the concept of substantial equivalence to identify any differences between the new food and its conventional correspondent.
Assessing the  security of  a genetically modified food raw-material, or its equivalence to conventional food, it is recommended that four elements are analyzed, namely:
(1) Parental variety, i.e. the plant originating the new genetically modified raw-material;
(2) Transformation process, including a characterization of the construct used  and the resulting event;
(3) Product of the inserted gene and potential toxicity and allergenicity and, finally;
(4) Composition of the new variety resulting from genetic transformation.
The data set of such analyses  shall enable identifying and characterizing any potential adverse effect associated with consumption of the new raw-material,  providing information to the risk management and risk communication phases.
Changes to  NK603 corn were caused by introducing the gene cp4 epsps, granting tolerance to the presence of the herbicide glyphosate in the pos-emergence phase of the plant. The engineered gene of the  EPSPS synthase is the most  studied transformation in plants, mainly soy and corn, with a large technical and scientific bibliography covering different aspects resulting from such transformation.
The organism donor of gene cp4 epsps was the  soil bacterium Agrobacterium sp., CP4strain. This organism donor of the gen inserted in NK603 corn is not used in the production of, nor used as, food. Besides, Agrobacteria species  are non pathogenic  to humans and animals and there are not reports that the epsps gene may be a determinant  of the pathogenicity   associated with Agrobacterium  in plants.
The EPSPS proteins catalyze the conversion of shikimic acid into 5-Enol-pyruvylshikimate-3-phosphate, which is an intermediary in the synthesis of aromatic amino acids and phenolic  compounds. For their functions, EPSPS are essential to normal growth of plants  and microorganisms. There is no toxicity associated to this family of proteins, which has a long history of environmental and nutritional safety. Besides, EPSPS proteins are not known for their persistence in the environment nor for affecting the phenotype of  the host organisms with  negative  properties, such as pathogenicity or potentially development into pests.
Several researchers conducted comprehensive characterizations of the CP4 EPSPS protein and the results showed that this protein has enzymatic properties equivalent to the EPSPS proteins endogenous to plants and microorganisms(14). In addition, detailed studies showed that the CP4 EPSPS protein is susceptive to proteolysis and enzymatic digestion, as it would be expected for an EPSPS  protein. Besides, the data available from acute oral toxicity studies, in vitro digestibility and bioinformatics comparison of CP4 EPSPS confirm its equivalence  to EPSPS proteins.
Toxicity tests were conducted with EPSPS-synthase isolated  from transformed plants. Harrison et.  al.(14) showed by means of ingestion by  gastric gavage that protein molecule  doses over one thousand  times the one found in modified seeds fail to cause cause physiologic  alterations in essayed animals. Results from in vitro proteolysis also confirm the prompt digestion and innocuousness of the engineered protein, removing any suspiciousness of allergenicity(14).
In addition to the history of safe use of  the  EPSPS  class proteins, studies were  conducted  that ratify the security of CP4 EPSPS proteins expressed in cultures such as soy, cotton, corn, canola and sugar beet. In one acute oral  toxicity study,  CP4 EPSPS was administered to  mice, in one high dose, to  confirm its safety(14). The results of such study showed that, as expected, the CP4 EPSPS protein is not toxic.
The CP4 EPSPS protein was produced and purified from E. coli, was characterized and showed to be equivalent  to the  CP4 EPSPS  produced in corn. The purified protein was orally administered to mice for acute toxicity assessment. Acute administration was deemed proper to assess the  security of CP4 EPSPS, since toxic proteins  act through acute mechanisms(23,18). The NOEL no observable effect for oral toxicity in mice was 572 mg/kg, the highest  tested dose(12). The result represented a  security margin of about two  hundred and sixty thousand (260,000) times, based on the average daily consumption of corn in the United States and expression of the protein in the glyphosate tolerant genetically modified  corn (assuming that there is no loss of  CP4 EPSPS during  processing). No statistically significant differences were observed in body weight, aggregate body weight or consumption of food among control groups (with the vehicle  or bovine albumin serum) and groups  treated with the purified CP4 EPSPS protein. This is an expected result, since  most EPSPS proteins are atoxic. The CP4  EPSPS protein showed to  be promptly digested in the gastric and  bowel fluids in vitro. Besides,  EPSPS proteins  have a safe history of use, since they are present common diet.
Dairy cows were fed with forage  and with glyphosate  tolerant genetically  modified corn, while milk production and composition was assessed  in  comparison with the  cows receiving the conventional plant(15). The results of this experiment indicate that there was no significant differences stemming from the food source, either in composition or yield of the milk produced by the animals.
Sheep fed with glyphosate tolerant canola were assessed regarding digestibility and permanence time of the recombinant DNA in the gastrointestinal  tract. The cp4 epsps gene and its fragments were monitored by PCR in real time and by conventional  amplification of intestinal fluids incubated at  39ºC during periods  of  time  from zero  to 240 minutes. Results showed  that genes  present  in fodder remained whole for a period relatively short, reducing  the likelihood of its absorption by the animal, and indicate also that intestinal microorganisms are responsible for the rapid degradation of DNA  at a pH 7(2).
One long term study was conducted in salmons, with diets based  on soybeans and corn modified with the cp4 epsps  gene(22). Upon completion of the essays, the authors verified that there  were no significant changes  in corporeal development, pyloric cecum and middle intestine of the  fish during the eight months of the experiment.
CP4 EPSPS proteins are ubiquitous in nature and  are  naturally present in foods derived from plant and microbial sources. The proteins  do not  show significant homology of  amino acids with proteins known to be toxic  or allergenic to mammals. Besides, the CP4 EPSPS protein was rapidly digested by in vitro gastric and bowel fluids. Further, EPSPS  proteins have a history of safe use and  are widely present  in  our diet.
Comparative allergenicity tests in sensibilized patients were  conducted with conventional soybean and corn, genetically modified and the respective isolated heterolog proteins(5). Products tested proved  to be  safe regarding their allergenic potential, since  there was no change in reactivity levels for modified kernel and  the isolated proteins failed  to cause  any reaction in sensibilized individuals.
Corn is not in the group of eight foods (milk, wheat, eggs,  fish, crustaceans, peanuts, soybean and nuts) that answer for about 90% of allergies in  humans. Allergy to corn may occur by ingestion of corn and its derivatives or through inhalation  of its flour or pollen. Corn has a long history in human and animal feeding, with rare cases of harm caused to health. According to Metcalfe (2003), allergy cases  reported in this  plant species are not  common, though recent studies show that previous diagnose methods may have underestimated the cases(21).
Several very consistent scientific studies proved that the nutritional value of NK603 Corn, or Roundup Ready Corn is, on average, equal  to that  of conventional corn. We emphasize “on average” because both corn types record variations (deviations from average), and this variation was shown to be similar in both cases. Given the specific characteristics of the NK603 corn production process, one may foresee that, when  cultivated under specific agronomic conditions (e.g., high competition with invading plants), the nutritional value of corn derived from such GMO is likely to be, in fact, higher than the conventional corn.
V. Environmental Aspects
Corn is a monoecious plant: a single individual has separately located male and female flowers. Corn plants are crossed fecundation plants and largely pollinated  with the help of wind, insects, gravity and others. Introduction of the genic elements mentioned above did not change the reproductive characteristics of the plant. Therefore, the likelihood  of crossed fecundation between hybrids and lineages of  non genetically modified  corn, and between plants of the NK603 event and other corn plants are the same.
Corn is the species that reached the highest domestication level among cultivated plants, and has lost its ability to  survive in nature such as, for instance, elimination of threshing. Thus, corn is a plant unable to survive in natural  conditions, without technical assistance. Therefore there is no possibility that corn changes into an invading plant or pest.
Genic flow in corn may take place through pollen transfer and through seed dispersion. Seed dispersion is easily controlled, since corn domestication eliminated  the  mechanisms for seed dispersion of the plant ancestors and the pollen movement is the only effective way for corn plant genes to escape.
Studies on corn pollen dispersion have been conducted, and some show that corn pollen may cover long distances. However, the majority of the released pollen is deposited close  to the culture, with a very low translocation rate outside the source culture. The main corn pollination agent is the wind and the distance that a viable pollen may cover depends on wind  pattern, moistness and temperature. Luna et. al.(19) assessed the isolation distance and control  of  pollen, and showed that crossed pollination happens in a distance up to  two hundred  meters  and no  crossed pollination took place in distances exceeding three hundred meters from the pollen sources, when the corn ear still keeps the silk. Results indicate that pollen viability is kept for  two hours and that crossed pollination was not observed in distances of three hundred meters from the pollen source.
With low  to  moderate winds,  estimates  are that, comparing  concentrations  at  1m from the source culture,  about 2% of  pollen are  recorded at 60m, 1.1% at 200m and 0,75-0,5% at 500m of distance. At 10m away from a field, on average, the number of pollen grains  per unit of area is ten times lower  than the figure recorded  at 1m from the border.  Therefore, if the  established separation distances developed for  production of corn  seeds  are  observed, one expects that the transference of pollen to  adjoining varieties is  minimized,  with no herbicide  tolerant generic material. Even in case of a genic escape, the probability of allele fixation containing the genic sequence conferring  tolerance to glyphosate in the population is very reduced  in the absence of selection pressure.
Cultivated corn is known for its interaction with different  organisms in the environment, including microorganisms, wild  animals  and soil and aerial invertebrates. Besides, corn is known for its susceptibility to different  fungi, viruses, bacteria, nematoids, pests caused by insects and acarine, use of pesticides  and other agricultural practices, such as  rotation of  cultures and use of  resistant  or tolerant genotypes developed by classic improvement. Interaction of  corn culture with wild vertebrates happen in large  number and is well known, since corn is  an excellent  source of nutrition. Such interactions happen with birds and mammals that live or find shelter in the agricultural environment  or  close to  this environment, in its borders, shrubs and dens. NK603 corn  has shown to  be equivalent  to conventional corn, except  for the glyphosate  tolerant characteristic. Its basic interactions with other organisms in the environment are not  held  as different from interactions of  conventional  corn. Though there is the potential  exposure  of corn culture weeds, pests and pathogens to CP4 EPSPS and CP4 EPSPS L214P proteins that are expressed in NK603 corn, there are no  concerns about the process causing adverse effects on such populations. Through trophic transference  and decomposition process,  organisms that are not targets of CP4 EPSPS and  CP4  EPSPS L214P proteins, such as predators  and  preys of corn  pests, may be exposed to very low levels of  the proteins without evidence of negative effects  on such organisms.
Environment safety of the EPSPS protein family is well accepted, since the proteins are ubiquitous in nature (bacteria, fungi, algae and higher plants), have no known toxicity, association with pathogenicity and do not grant comparative selective advantage to plants that contain the proteins. Though EPSPS  proteins are found in plants and microorganisms, primary amino acid sequences display considerable differences. Analyses of alignments using peptides (ALLPEPTIDES) show  that members of the EPSPS protein family may have less than 25% of common identity in a window  of  about 450 amino  acids (corresponding to  the total size of CP4 EPSPS). Despite this low identity level of amino acid sequences, EPSPS  family proteins are highly related in terms  of structure and function(12).
The EPSPS enzyme has no target organism, since it is an enzyme  involved in the biochemical way of shikimic acid  in plants and  microorganisms. Any non target organism  that interacts with a plant culture displays a close interaction with  a number of plants and microorganisms and therefore is constantly exposed to EPSPS family proteins. Based on the history of occurrence and safe exposure of this protein family, there is no evidence that any EPSPS protein  may display biological  activity on a non target  organism.
VI. Restrictions to the  use of the GMO and its derivatives
According to Article 1 of Law no. 11,460, of March 21, 2007 ”research and  cultivation of  genetically modified organisms may not be conducted in  Amerindian areas and conservation units”.
Technical opinions related  to agronomic  performance reached a  conclusion that there  is equivalence between transgenic  and  conventional  plants. Thus, this information suggest  that transgenic  plants are not fundamentally different from the non-transformed corn genotypes, except  for  the  tolerance to glyphosate. Besides, there is no evidence of adverse reactions to the use of NK603 corn. For this reason, there are  no restrictions to the use of this corn or its derivatives, either for human  or animal feeding.
The vertical  genic  flow to  local varieties (the  so-called  Creole corns)  of  open pollination is possible and carries the  same  risk caused by the commercial genotypes  available in the market (80% of the conventional corn farmed in Brazil come from commercial seeds that underwent a process of genetic improvement). Coexistence among cultivars of conventional (either cultivated of local varieties) and transgenic cultivars is possible  from the agronomic viewpoint(6,20).
After ten years of use in different countries, there was no problem  detected  for human and animal health, or to the environment that  may be ascribed to transgenic maize. It shall be emphasized that the lack of negative  results in cultivation of transgenic corn plants does not mean that this may not  happen. Zero risk coupled with absolute safety is an inexistent  combination in the  biologic world, although there is a host  of trustworthy scientific information and a safe historic of  use of ten years that enable us to argue that  the  NK603 corn is as  safe as conventional versions. Therefore, the applicant shall conduct post-commercial release monitoring under CTNBio Ruling Instruction no. 3.
VII. Considerations on particulars of different regions of  the  country (information to supervising bodies)
In Brazil, there are no kindred  corn species in natural distribution.
VIII. Conclusion
Whereas:
1. Corn is the species that reached  the highest domestication level among cultivated plants, and is  unable to survive  in nature with no human intervention.
2. In Brazil, there are no wild species  with which corn may intercross, since  the closest wild corn species is teosinte, found only in Mexico and in some Central America locations, where it may cross  with corn cultivated in production fields.
3. There is established knowledge on safety of the use  of  corn in the human and animal food chain.
4. The transformation event in analysis failed to modify the composition and the nutritional value of corn.
5. There is no evidence that the  transgene or the transformation event may cause adverse effects to human and animal health.
6. The history of NK603 corn use in the European Union (since 1999), USA (since 2000) and Canada (since 2001), in addition to the data  on field  tests  conducted in Brazil (since 2000) give  sufficient evidence that the kernel and products derived from Glyphosate Tolerant NK603 Corn are as safe as those of conventional corn.
7. There is robust scientific evidence that NK603 corn has no adverse  effects  on human and animal health, and this  information is based in international scientific literature.
8. In 2003, the European Food Safety Authority (EFSA) examined a request for commercial  release of NK603 corn and  issue  a favorable scientific opinion on December 4, 2003, with  a conclusion that “NK603 corn is as safe as conventional corn and therefore the marketing of NK603 for  processing and its use in human and animal food  are unlikely to have any adverse effect on human and animal health or, in this context, on the environment.”
9. Given the detailed data provided  by the applicant, the results obtained in control and security essays  of the genetically modified organism in analysis, the  elements credited to  authors of scientific work mentioned  and  inexistence of evidence contrary to nutritional, toxicological  and  allergenic security, after  thorough investigation, we  are favorable to  commercial release of  NK603 corn for consumption in the human and animal food  chain.
10. Corn is a plant that is unable to survive in natural conditions, without technical assistance. This  way, there  is  no likelihood that corn may  transform into  an invading plant or weed.
11. Environmental  safety of the  EPSPS family proteins  is well accepted, since the  proteins are ubiquitous in nature (bacteria, fungi, algae and higher plants), exhibit no known toxicity, are  not associated  to pathogenicity and  fail  to grant any selective advantage  to plants that do not contain such proteins.
12.  In case of a genic  escape, the likelihood of fixation of an  allele containing the genic sequence that grants tolerance  to glyphosate  in the  population is minimal in absence of selection pressure.
13. Basic  interactions of NK603 corn  with other organisms in the environment are not  held  different from interactions of conventional corn.
14. Organisms  that are not targets of CP4 EPSPS and CP4  EPSPS L214P, such as predators  and preys of corn pests, may be exposed to  very low levels  of  the proteins without, however, evidence  of negative effects on such organisms.
15. There is no evidence that any EPSPS  protein  will have  biologic activity on  non-target organisms.
For the foregoing, and  considering internationally accepted criteria in the process of  risk analysis  for  genetically modified raw-materials, a conclusion  emerges that Roundup  Ready  corn, derived from NK603, is as safe as  its conventional  equivalent. Third party independent scientific studies and publications provided by the applicant were also taken into consideration and  consulted.
CTNBio holds  that the activity is not  a potential cause  of significant degradation to the environment nor of  harm to  human and animal health. Restrictions to the use of the GMO analyzed and  its  derivatives  are conditioned to the provisions  of CTNBio Ruling Resolutions no. 03 and 04.
CTNBio analysis took into consideration opinions issued by  members of the Commission and ad hoc consultants; documents delivered by  applicant to CTNBio Office of the Executive Secretary; results from planned releases to the  environment; lectures, texts and discussion of the  public hearing held on 03.20.2007. Also taken into consideration were applicant’s studies and publications, conducted  by third parties.
According to Annex I of  Ruling Resolution no. 5, of March  12, 2008, the applicant shall have thirty (30) days from the publication of this  Technical Opinion to harmonize its  proposal for post-commercial release monitoring.
IX. Bibliography mentioned
1. AGBIOS. GM Database. 2008. Biotech Crop Database. Corn MON NK603. http://wwww.agbios.com
2. ALEXANDER T.W.; SHARMA R.; DENG M.Y; WHETSELL A.J.; JENNINGS J.C.; WANG Y.; OKINE E. DAMGAARD D.; McALLISTER T.A. 2004. Use of quantitative real-time and conventional PCR to assess the stability of the cp4 epsps transgene from Roundup Ready® canola in the intestinal, ruminal, and fecal contents of sheep. J. Biotechnol. 112: 255-266.
3. ARONSON A.I. 2000. Incorporation of protease k into larval insect membrane vesicles does not result in disruption of integrity or function of the pore-forming bacillus thuringiensis delta-endotoxin. Appl. Environ. Biol. 66: 4568-4570.
4. BAHIA FILHO A.F.C.; GARCIA J.C. 2000. Análise e avaliação do Mercado brasileiro de sementes de milho. In: UDY C.V.; DUARTE W.F. (Org.) Uma história brasileira do milho: O valor de recursos genéticos. Brasília:  Paralelo 15. 167-172.
5. BATISTA R.; NUNES B.; CARMO M.; CARDOSO C.; JOSÉ H.; DE ALMEIDA A.; MANIQUE A.; BENTO L.; RICARDO C.; OLIVEIRA M. 2005 Lack of detectable allergenicity of transgenic maize and soya samples.J. allergy Clin. Immunol. 116: 403-10,2005.
6. BROOKES G.; BARFOOT P.; MELÉ E.; MESSEGUER J.; BÉNÉTRIX F. BLOC D.; FOUEILLASSAR C; FABIÉ A.; POEYDOMENGE C. 2004. Genetically modified maize: pollen movement and crop co-existence. Dorchester, UK: PG economics, 20pp (www.pgeconomics.co.uk/pdf/Maizepollennov2004final.pdf)
7. CARPENTER J.; FELSOT A; GOOGE T.; HAMMING M; ONSTAD D. E SANKULA S. 2002. Comparative environmental impacts of biotechnology-derived and traditional soybean, corn and cotton crops. Council for Agricultural and Science Technology Report, Ames, Iowa. 200p. Available at: www.cast-science.org. Sponsored by the United Soybean Board. www.unitedsoybean.org. (annex 1-2)
8. CONAB. Milho total (1ª e 2ª safra) Brasil – Série histórica de area plantada – safra 1976-77 a 2006-07. Disponível em http://www.conab.gov.br/download/safra/MilhoTotalSerieHist.xls
9. FABO/WHO – Food and agriculture Organization of the United Nations / World Health Organisation 2000. Grassland Index. Zea mays L. (http://www.fao.org/WAICENT/faoinfo/agricult/agp/agpc/doc/gbase/data/pf000342.htm)
10. FAO. 2007. FAOSTAT. Disponível em: http://faostat.fao.org.site.340/default.aspx.
11. FAOSTAT-Agriculture – Food And Agriculture Organization Of The United Nations – Disponível  em: http://faostat.fao.org.site.340/default.aspx. Hain R & Schreier PH (1996) Genetic engineering in crop protection: opportunities, risks
12. FUCHS R.L. 1998. Assessment of the allergenic potential of foods derived from genetically engineered plants: glyphosate tolerant soybean as a case study. In: EISENBRAND G.; AULEPP H.; DAYAN M. (Eds.) Food Allergies And Intolerances Symposium. Weinheim: VCH Verlagsgesellschaft mbH, 17:212-221.
13. GIANESSI L.; SILVERS C.; SANKULA, S E CARPENTER J. 2002. Executive Summary-Plant Biotechnology – Current and Potential Impact for Improving Pest Management in Us Agriculture. An Analysis OF Case Studies. NCFAP. National Center for food and Agricultural Policy: 1-23. Disponível em http//www.ncfap.org/40caseStudies.htm
14. HARRISON L.A.; BAILEY M.R.; NAYLOR M.W.; REAM J.E.; HAMMOND B.G.; NIDA G.L.; BURNETTE B.L.; NICKSON T.E.; MITSKY T.A.; TAYLOR M.L.; FUCHS R.L.; PADGETTES.R. 1996. The expressed protein in glyphosate synthase from Agrobacterium sp. Strain CP4, is rapidly digested in vitro and is not toxic to acutely gavaged mice. J. Nutr. 126: 728-740.
15. IPHARRAGUERRE I.R.; YOUNKER R.S.; CLARK J.H. 2003 Performance of lactating dairy cows fed corn as whole plant silage and grain produced from a glyphosate-tolerant hybrids (event NK 603). J. Dairy Sci. 86: 1734-1741.
16. JOHNSON W.G.; BRADLEY P.R.; HART S.E.; BUESINGER M;L E MASSEY R.E.2000. Efficacy and economics of weed management in glyphosate-resistant corn (Zea mays). Weed Technol. 14: 57-65.
17. KLEIN T.M.; WOLF E.D.; WU R.; SANFORD J.C. 1987. High velocity microprojectiles for delivering nucleic acids into living cells. Nature 327:70-73.
18. KESSLER D.A.; TAYLOR M.R.; MARYANSKI J.H.; FLAMM E.L.; KAHL, L.S. 1992. The safety of foods developed by biotechnology. Science 256: 1747-1749.
19. LUNA S.V.; FIGUEROA J.M; BALTAZAR B.; GOMEZ L.T.; TOWNSEND R. E SCHOPER J.B. 2001. Maize pollen longevity and distance isolation requirements for effective pollen control. Crop Sci. 41: 1551-1557.
20. MESSEGUER J.; PEÑAS G.; BALLESTER J.; BAS M.; SERRA J. SALVIA J.; PAULADEMÁS M.; MELÉ E. 2006. Pollen-mediated gene flow in maize in real situations of coexistence. Plant Biotechnology Journal. 4: 633-645.
21. METCALFE D.D.2003. Introduction: What are the issues in addressing  the allergenic potential of genetically modified foods? Environ. Health Perspect. 111: 1110-1113.
22. SANDEN M.; BERNTSSEN M.H.G..; KROGDAHL. Å.; HEMRE G.-I.; BAKKE-McKELLEP A.-M. 2005. An examination of the intestinal tract of Atlantic salmon, Salmo salar L., parr fed different varieties of soy and maize. J. Fish Dis. 28: 317-330.
23. SJOBLAD R.D.; MCCLINTOCK J.T.; ENGLER R. 1992. Toxicological considerations for protein components of biological pesticide products. Regul Toxicol Pharmacol. 15: 3-9.
24. SPALHANET. Descontrole: Uso de Agrotóxico é o Principal Problemas de Saúde dos Agricultores. 2006. Ano XXI N° 164. www.sinpaf.org.br
25. THARP B.E.; KELLS J.J. 1999. Influence of herbicide application rate, timing, and interrow cultivation on weed control and corn (Zea mays). Yield in glufosinate-resistance and glyphosate-resistant corn. Weed Technol. 13: 807-813.
X. Bibliography consulted
1. ALTIERI M. A. 2005. The myth of coexistence: Why transgenic crops are not compatible with agroecologically based systems of production. Bull. Sci. Technol. & Soc. 25: 361-371.
2. AUMAITRE A.; AULRICH K.; CHESSON A.; FLACHOWSKY G.; PIVA G. 2002. New feeds from genetically modified plants: substantial equivalence, nutritional equivalence, digestibility, and safety for animals and the food chain. Livestock Prod. Sci. 74: 223-238.
3. BENBROOCK C. 2003. Impacts of genetically engineered crops on pesticide use in United States: The first eight years. Biotech Infonet Paper No., www.biotechinfo.net/technicalpapaer6.html.
4. BENEDITO V.A.; FIGUEIRA A. V. 2008. Segurança ambiental. In: Biotecnologia e Meio Ambiente. BOREM A.; GIUDICE M.; DEL P. p. 167-198.
5. BOREM A. 2001. Escape gênico & transgênicos. Viçosa, UFV 206 p.
6. BRINGOLF R.B.; COPE W.G.; MOSHER S.; BARNHART M.C.; SHEA D. 2007. Acute and chronic toxicity of glyphosate compounds to glochidia and juveniles of Lampsilis siliquoidea (Unionidae). Environ. Toxicol. Chem. 26: 2094-100.
7. BROOKES G. et al. 2004. GM Maize – Pollen Movement and Crop-existence. Dorchester, UK: PG Economics Ltda. http://www.pgeconomics.co.uk
8. BROOKES G.; BARFOOT P. 2006. Global Impact of Biotech Crops: Socio-Economic and Environmental Effects  in the First Ten Years of Commercial Use. AgBioforum. 9: 139.151.
9. CHRENKOVA M.; SOMMER A.; CERESNAKOVA Z.; NITRAYOVA S.; PROSTREDNA M. 2002. Nutritional evaluation of genetically modified maize corn performed on rats. Archives of Animal Nutrition – Archiv Fur Tierernahrung 56: 229-235.
10. CODEX ALIMENTARIUS. 2003. Guideline for the conduct of Food safety assessment of food derived from recombinant-DNA plants. the codex Alimentarius Commission and the FAO/WHO Food standards Programme. Rome. CAC/GL 45-2003. ftp://ftp.fao.org/es/esu/food_guide_plants_en.pdf
11. COE JR. E.H. 2001. The origin of maize genetics. Nature Rev. Genetics 2:898-906.
12. CONNER A.; GLARE T.E.; NAP J-P. 2003. The release of genetically modified crops into the environment. Part II – Overview of ecological risk assessment. The Plant J. 33:19-46.
13. DEMANÈCHE S.; SANGUIN H.; POTÉ J.; NAVARRO E.; BERNILLON D.; MAVINGUI P.; WILDI W.; VOGEL T.M.; SIMONET P. 2008. Antibiotic-resistant soil bacteria in transgenic plant  fields. Proc. Natl. Acad. Sci. USA. 105: 3957-3962.
14. DOMINGO J.L. 2000. Health Risks of GM foods.: Many opinions but few data. Science 288: 1748-1749.
15. EASTHAM K.; SWEET J. 2002. Genetically modified organisms (GMO): the significance of gene flow through pollen transfer. Copenhagen: European Environmental Agency.
16. DONKIN S.S.; VELEZ J.C; TOTTEN A.K; et al. 2003. Effects of feeding silage and grain from glyphosate-tolerant or insect-protected corn hybrids on feed intake, ruminal digestion, and milk production in dairy cattle. J. Dairy Sci. 86: 1780-1788.
17. DOUVILLEA M.; GAGNÉ F; ANDRÉA C.; BLAISEA C. 2008. Ocurrence of the transgenic corn cry1Ab gene in freshwater mussels (Elliptio companata) near corn fields: Evidence of Exposure by bacterial ingestion. Ecotoxicol. Environ. Saf. (in press).
18. EMBERLIN J. 1999. The dispersal of corn pollen Zea mays: A report based on evidence available from publications and internet sites. National Pollen Research Unit, University College, Worcester WR2 AJ, United Kingdom.
19. ERICKSON G.E.; ROBBINS N.D.; SIMON J.J.; et al. Effect of feeding glyphosate-tolerant (Roundup-Ready events GA21 or NK603) corn compared with reference hybrids on feedlot steer performance and carcass characteristics. J. An. Sci. 81: 2600-2608.
20. EUROPEAN COMMISSION. 2006. Technical Report EUR 22102 EM. New Case Studies on the Coexistence of GM and Non-GM Crops in European Agriculture. http://www.jrc.es/home/pages/eur22102enfinal.pdf
21. EUROPEAN FOOD SAFETY AUTHORITY (EFSA). 2006. Guidance document of the scientific panel on genetically modified organisms for risk assessment of genetically modified plants and derived food and feed. EFSA J. 99:1-100.
22. FOOD AND AGRICULTURE ORGANIZATION OF THE UNITED NATIONS . 2004. The State of Food and Agriculture. Agriculture Biotechnology. Meeting the Poor. Rome, 208pp. http://www.fao.org
23. FOOD SAFETY AND GMOS. 2004. Consensus Document. 10pp. http://www.cedab.it
24. FORSBACH A.; SCHUBERT D.; LECHTENBERG B.; GILS M.; SHIMIDT R. 2003. A Comprehensive characterization of single-copy T-DNA insertion in the Arabidopsis thaliana genome plant Mol. Biol. 52:161-176.
25. GARCIA M. A.; ALTIERI M. A. 2005. Transgenic crops: implications for biodiversity and sustainable agriculture. Bull. Sci. Technol. & Soc. 25: 335-353.
26. GELMINI G.A.; VICTORIA FILHO R.; NOVO M.C.S.S.; ADORYAN M.L. 2005. Resistência de Euphorbia heterophylla L. aos herbicidas inibidores das ALS na cultura da soja. Sci. Agric. 62: 452-457.
27. GERTZ JR. J.M.; VENCIL W.K.; HILL N.S.1999.Tolerance of transgenic soybean (Glycine max) to heat stress. In: Proceedings of the 1999 Brighton Conference Weeds ( The BCPC Conference). Vol. 3.; November 1999; Brighton, UK, p. 835.840
28. GIESY J.P.; DODSON S.; SOLOMON K.R. 2000. Ecotoxicological risk assessment for Roundup herbicide. Rev. Environ, Contam. Toxicol. 167: 35-120
29. GIULIETTI A.M.; HARLEY R.M.; QUEIROZ L.P; WANDERLEY M.G.L.; VAN DEN BERG C. 2005. Biodiversidade e conservação das plantas no Brasil. Megadiv. 1: 52-61.
30. GLADYSHEV E.A.; MESELSON M.; ARKHIPVA I.R.2008. Massive horizontal gene transfer in bdelloid rotifers. Science. 320: 1210-1213.
31. GLIESSMAN S.R. 2005. Agroecologia: processos ecológicos em agricultura sustentável. Porto Alegre: UFRGS. 652P.
32. HAMMOND B.G.; VICINI J.L. HARTNELL G.F.; NAYLOR M.W. KNIGHT C.D.; ROBINSON E.H.; FUCHS R.L.; PASGETTE S.R. 1996. The feeding value of soybeans fed to rats, chickens, catfish and dairy cattle is not altered by incorporation of glyphosate tolerance. J. Nutrs. 126: 717-727.
33. ICOZ I.; SAXENA D.; ANDOW D.A.; ZWAHLEN C.; STOTZKY G. 2008. Microbial populations and enzyme activities in soil in situ under transgenic corn expressing cry proteins from Bacillus thuringiensis. J. Environ. Qual. 37: 647-62.
34. JAMES C. 2006. Global status of commercialized Biotech/GM Crops. ISAAA Brief 35. ISAAA: Ithaca Ny. Website: http://www.isaaa.org
35. KAY R.; CHAN A.; MCPHERSON J. 1985. Duplication of CaMV 35S promoter sequences creates a strong enhancer for plant genes. Science 236: 1299-1302
36. KINUPP V.; BARROS I.B.I. 2004. Levantamento de dados e divulgação do potencial das plantas alimentícias alternativas no Brasil. Horticultura brasileira V.4.
37. KINUPP V. 2007. Plants alimentícias não convencionais na Região Metropolitana de Porto Alegre. Programa de Pós-Graduação em Fitotecnia – Faculdade de Agronomia –UFRGS. 590p.
38. KLEE H.J; MUSKOPF Y.M.; GASSER C.S. 1897. Cloning of an Arabidopsis thaliana gene encoding 5-enolpyruvylshikimate-3-phosphate synthase: sequence analysis and manipulation to obtain glyphosate-tolerant plants. Mol Gen Genet. 210: 437-442.
39. LATHAM J.R.; WILSON A.K.; STEINBRECHER R.A. 2006. The mutational consequences of plant transformation. J. of Biomed. Biotech. 2006: 1-7.
40. MALATESTA et. Al. 2002. Ultrastructural morphometrical and immunocytochemical of hepatocytenuclei from mice fed on genetically modified soybean. Cell structure and function. 27:173-180.
41. MESSEGUER J.; PEÑAS G.; BALLESTER J.; BAS M.; SERRA J.; PALAUDELMÀS M.; MELÉ E. 2006. Pollen-mediated gene flow in corn in real situations of coexistence. Plant Biotechnology Journal. 4:633-645.
42. MONIER J.M.; BERNILLON D.; KAY E.; FAUGIER A.; RYBALKA O.; DESSAUX Y.; SIMONET P.; VOGEL T.M. 2007. Detection of potential transgenic plant DNA recipients among soil bacteria. Environ Biosaf. Res. 6:71-83.
43. MONQUERO P.A; SILVA A.C. 2007. Efeito do período de chuva no controle Euphorbia heterophylla e Ipomea purpurea pelos herbicidas glyphosate e sulfotase. Planta Daninha 25:399-404.
44. MOREIRA M.S.; NICOLAI M.; CARVALHO S.J.P; CHRISTOFFOLETI P.J. 2007. Resistência de Conyza Canadensis e C. bonariensis ao herbicida glyphosate. Planta Daninha. 25: 157-164.
45. NIELSEN K.M.; BONES A.M.; SMALLA K.; VAN ELSAS J.D. 1998. horizontal gene transfer from transgenic plants to terrestrial bacterial – a rare avent? FEMS microbial Rev. 22: 79-103.
46. OECD. 2003. Considerations for the safety assessment of animal feedstuffs derived from genetically modified plants. Series on the safety of Novel Foods and Feeds, No. 9. Organization for Economical Co-operation and Development. ENV/JM/MONO ,10.46p.
47. OFFICIAL JOURNAL OF THE EUROPEAN UNION. http://www.agbios.com/docroot/decdocs/06-286.015pdf
48. PADGETTE S.R.; KOLACZ K.H.; DELANNAY X.; RE D.B.; LaVALLE B.J.; TINIUS C.N.; RHODES W.K.; OTERO Y.I.; BARRY G.F.; EICHHOLTZ D.A.; PESCHKE V.M.; NIDA D.L.; TAYLOR N.B; KISHORE G.M. 1995. Development, identification and characterization of a glyphosate-tolerant soybean line. Crop Sci. 35: 1451-1461.
49. PADGETTE S.R.; TAYLOR N.B.; NIVIA D.L.; BAULEY M.R.; MacDONALD J.; HOLDEN L.R.; FUCHS R.L. 1996. The composition of glyphosate-tolerant soybean seeds is equivalent to that of conventional soybeans. J. Nutr. 16:702.716.
50. PONTIROLI A.; SIMONET P.; FROSTEGARD A.; VOGEL T.M.; MONIVER J.M. 2007. Fate of transgenic plant DNA in the environment, Environ. Biosaf. Res. 6: 15-35.
51. PRAY C.E.; HUANG J. 2003. The Impact of Bt Cotton in China. In: Economic and Environmental Impacts of Agbiotech: A Global Perspective. KALAITZANDONAKES N. (Ed.). New York: Kluwer-Plenum Academic Publisher.
52. RAPOPORT E. H.; LADIO A.; RAFFAELE E.; GHERMANDI L.; SANZ E.H. 1998. Malezas Comestíbles. Hay yuyos y yuyos. Ciencia Hoy, v.9. n.49.
53. RAPOPORT E.H.; DRAUSAL B.S. 2001. Edible plants, In: Encyclopedia of biodiversity. LEVIN S. (ed). New York: Academic Press, 375-382.
54. SABHARWAL N.; ICOZ I., SAXENA D.; STOSZKY G. 2007. Release of the recombinant proteins, human serum albumin, beta-glucuronidase, glycoprotein B from human cytomegalovirus, and green fluorescent protein, in root exudates from transgenic tobacco and their effects on microbes and enzymatic activities in soil. Plant Physiol Biochem. 45: 464-469.
55. SAXENA D.; FLORES S.; STOTZKY G. 1999. Insecticidal toxin in root exudates from Bt corn. Nature 402: 480.
56. SHAN G.; EMBREY S.K.; HERMAN R.A.; MCCORMICK R. 2008. Cry1F protein not detected in soil after three years of transgenic Bt corn (1507 corn) use. Environ Entomol. 37:255-62.
57. SHEWRY P.R.; et al.; 2007, Are GM and conventionally Bred Cereals Really Different? Food Sci. Technol. 18: 201.209.
58. SIQUEIRA J.O.; TRANNIN. 2008. Agrossistemas transgênicos. In: Biotecnologia e Meio Ambiente. Borem A.; Giudice M. DEL P. p. 225-309.
59. TAYLOR M.L.; HARTNELL G.F.; RIORDAN S.G.; et al. 2004. Comparison of broiler performance when fed diets containing grain from Roundup Ready (NK603), YieldGard x Roundup Ready (MON 810 X NK603), non-transgenic control, or commercial corn, Poult. Sci. 83: 1758-1758.
60. TAYLOR M.L.; HARTNELL G.; NEMETH M.; KARUNANANDAA K.; GEORGE B. 2005. Comparison of broiler performance when fed diets containing corn grain with insect-protected (corn rootworm and European corn borer) and herbicide-tolerant (glyphosate) traits, control corn, or commercial reference corn-revisted. Poult. Sci. 84: 1493-1899.
61. NUFFIELD COUNCIL ON BIOETHICS. 2004. The use of genetically modified crops in developing countries. 122pp. http://www.nuffieldbioethics.org
62. TRAVIK T.; HEINEMANN J. 2007. Genetic Engineering and Omitted Health Research: Still No answers to ageing questions. Third Word Network, 36p.
63. U.S. FOOD AND DRUG ADMINISTRATION. 2006. Recommendations for the early food safety evaluation of new-pesticidal proteins produced by new plant varieties intended for food use. Center for Food Safety and Applied Nutrition. U.S. Food and Drug Administration. http://www.cfsan.fda.gov/~dms/bioprgu2.htmal
64. VARGAS L.; BIANCHI M.A.; RIZZARDI M.A.; AGOSTINETTO D.; DAL MAGRO T. Conyza bonariensis resiste ao glyphosate na região sul do Brasil. Planta Daninha 25: 573.578.
65. VARGAS L.; RIZZARDI M.A.; BIANCHIM A.; 2007. Manejo e controle de espécies tolerantes ou resistentes ao glifosato. Plantio Direto mar/abr. 2007: p.30-33
66. VIDAL R.A; TREZZI M.M.; STACHLER J., LOUX M. 2007. Definindo resistência aos herbicidas. Plantio Direto jul/ago.2007.
67. VIDAL R.A.Q.; LAMEGO F.P. TREZZI M.M. 2006. Diagnóstico da resistência aos herbicidas em plantas daninhas. Planta Daninha 24: 597-604.
68. WILLIAMS G.M.; KROES R.; MUNRO I.C. 2000. Safety evaluation and risk assessment of the herbicide Roundup and its active ingredient, glyphosate, for humans. Regul. Toxicol. Pharmacol. 31: 117-165.
69. WORD HEALTH ORGANIZATION (WHO). 2005. Modern Food Biotechnology, Hunan Health and Development: An Evidence-Based Study. 76pp. http//www.who.int/foodsafety
70. YASMIN S.; D’SOUZA D. 2007. Effect of pesticides on the reproductive output of Eisena Fetida. Bull. Environ, Contam. Toxical. 79: 529-532.

Walter Colli
President  of CTNBio

Dissenting vote:
The author of the Technical  Opinion, Dr. Paulo Brack  (Permanent Environment Sectoral Subcommission) issued an opinion contrary to approval  of this product for  the following reasons, verbatim:
1. Aspects linked to biodiversity  are fundamental in analyzing and debating the prospective commercial release of transgenic varieties, since genetically modified plant technology should not be part of the usual procedure of increasing impoverishment of  the rich biodiversity  stock and decay of ecologic  resilience and self-regulation processes.
2. Increased resistance to the glyphosate herbicide may occur, as already happened with soybeans, increasing the  invasion by adventitious plants, since  the herbicide may become innocuous and fail to bring economic  advantages.
3. There are few experimental studies regarding environmental effects of this technology over the environment.
4. Cost-benefit  analyses of GMO should be coupled  to  a wider, or systemic, assessment of causes of  maladjustments to which these organisms have been created and placed in the market.
5. The absence of works, in Brazil, on the possible effect of transgenic  plants on  the dynamic changes of soil microorganisms.
6. There is evidence  that  relatively large DNA fragments of  GM  plants do survive for large periods after digestion, and are detectable in feces.
7. There are still  large knowledge gaps regarding bio-risks  of an open pollination plant that represents a very important culture for the  small farmer and for human and animal feeding.
8 There  are few studies on the distance the pollen may reach and the likelihood that the gynaeceum is reached. This aspect was not taken into  account until now, and there are not studies on the presence of other pollinators, such as wild  bees in the different Brazilian biomes.
9. Despite the anemophily strategy of the plant, by the  abundant production of pollen  in staminate  flowers, the risk of genic flow  between  GM and non-GM corn by Apis mellifera L. may  exist.
10. In Brazil there is a possible contamination, regarding  GM  corn, mainly because of absence of an efficient mechanism for seed segregation.
11. There are no programs to protect the producer against contamination of seed or corn kernel production.
12. A GM plant seed may propagate indefinitely and, in case problems  occur, there is no  way to  known whether collection will take place.
13. There are no detailed studies  on the question of stability of locations,  the  sites of current  insertion, the number and stability of  inserts, the effects of the transgenic promoter, the patterns of  inserts and mutations post-genetic modifications in the coded protein and  regulation sequence  under conditions  beyond experiments confined to  laboratories.
14. Unpredictable effects in  the GMO genetic stability may cause changes  to nutritional value or even allergenicity or other inconvenient factor  to human health.
15. There is  no conclusive studies conducted by the applicant on toxicological and allergenic aspects of  the event  to be analyzed.
16. Aspects related  to system diversity shall be part of the studies, including the likelihood that there is no genetic erosion and even larger biodiversity loss.
17. There was not an assessment of environmental risk nor environmental impact studies.
18. There is no previous studies of such plants and possible environmental consequences in Brazilian ecosystems.
19. Risks for other “non-target” organisms are unknown (bees, birds, soil microorganisms resulting from degradation of modified plants, etc.)
20. The Cartagena Protocol  on Biosafety is little considered in Brazil, mainly in what is established in Annex II thereof.
21. The traditional practice of interchanging seeds among farmers