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In a submission dated October 13, 2006, Monsanto Company (Monsanto) provided to the Food and Drug Administration (FDA) a summary of the safety and nutritional assessment of a new bioengineered insect-resistant corn line, corn event MON 89034. Monsanto provided additional information on January 18 and March 8, 2007. Monsanto concluded that food and feed derived from the new insect-resistant corn is as safe and nutritious as food and feed derived from conventional corn varieties currently being marketed.
The intended effect of the modification in corn event MON 89034 is to confer resistance to a broad spectrum of Lepidopteran insects. To accomplish this objective, Monsanto introduced the chimeric cry1A.105 gene (derived from Bacillus thuringiensis (B.t) subsp. aizawai and B.t. subsp. kurstaki) and the cry2Ab2 gene (derived from B. t. subsp. kurstaki) into an inbred conventional corn line LH172 and designated the resulting corn event as MON 89034. The Cry1A.105 and Cry2Ab2 proteins1 encoded by these genes confer resistance to Lepidopteran insects, including European corn borer, Asian corn borer, southwestern corn borer, and sugarcane borer.
The Environmental Protection Agency (EPA) regulates plant-incorporated protectants under the Federal Food, Drug, and Cosmetic Act and the Federal Insecticide, Fungicide, and Rodenticide Act. Under EPA regulations, the Cry1A.105 and Cry2Ab2 proteins in corn event MON 89034 are considered pesticidal substances. Therefore, the safety assessment of these proteins falls under the regulatory purview of EPA.
To generate the corn event MON 89034, Monsanto used an Agrobacterium tumefaciens-mediated transformation system. Monsanto transformed the conventional corn line using the binary plasmid vector PV-ZMIR245. This vector contains two separate T-DNAs. The first T-DNA (designated T-DNA I) contains the cry1A.105 and the cry2Ab2 expression cassettes. The second T-DNA (designated T-DNA II) contains the nptII expression cassette encoding the neomycin phosphotransferase enzyme which confers tolerance to aminoglycoside antibiotics and which was used as a selectable marker in the initial screening of transformants. During transformation, both T-DNAs were inserted into the plant genome. Once the transformants were identified, Monsanto used traditional breeding to select plants containing T-DNA I but not T-DNA II. The T-DNA I region and T-DNA II region contain the elements listed in Table 1 in addition to Left and Right Border sequences and intervening sequences.
| Genetic element in T-DNA I | Description |
|---|---|
| P-e35S | The promoter and leader from the Cauliflower Mosaic Virus (CaMV) 35S RNA containing the duplicated enhancer region |
| L-Cab | 5' untranslated leader of wheat chlorophyll a/b-binding protein |
| I-Ract1 | Intron from rice actin gene |
| CS-cry1A.105 | Coding sequence for B.t. Cry1A.105 protein |
| T-Hsp17 | 3' nontranslated region of the coding sequence for wheat heat shock protein 17.3, which ends transcription and directs polyadenylation |
| P-FMV | 35S promoter from Figwort Mosaic Virus |
| I-Hsp70 | First intron from maize heat shock protein 70 gene. |
| TS-SSU-CTP | DNA region containing the targeting sequence for the transit peptide region of maize ribulose 1,5-biphosphate carboxylase small subunit and first intron |
| cry2Ab2 | Coding sequence for B.t. Cry2Ab2 protein |
| T-nos | Terminator from A. tumefaciens |
| Genetic element in T-DNA II | Description |
|---|---|
| T-nos | Terminator from A. tumefaciens |
| CS-nptII | Coding sequence for neomycin phosphotransferase II |
| P-35S | The promoter and leader from CaMV 35S RNA |
Monsanto used restriction enzyme digestion of genomic DNA followed by Southern blot analysis to characterize the inserted DNA in MON 89034. Monsanto states that based on the results obtained from these analyses, the T-DNA I from plasmid PV-ZMIR245 is present in MON 89034 as a single functional copy. Monsanto reports that T-DNA II sequences including nptII sequences were not detected in MON 89034. Monsanto further states that the insertion in MON 89034 was stable within and across multiple (seven) generations. Monsanto reports some molecular rearrangement of the e35S promoter which regulates expression of the cry1A.105 gene and replacement of the Right Border sequence with a Left Border sequence but notes that these changes are not critical for overall protein expression. Monsanto also assessed Mendelian segregation of event MON 89034 and concluded that T-DNA I containing the cry1A.105 and cry2Ab2 cassettes is inherited as a single dominant trait.
Corn grain (kernel) and its processed fractions are consumed as human food and animal feed. The majority of corn is used as animal feed; the remainder is exported, held as ending stock, processed into corn syrup, converted to ethanol, extracted for starch, used as processed food, and grown as corn seed. The kernel consists of pericarp, germ, and endosperm. The pericarp can be removed during processing to yield the germ (containing 50% oil) and endosperm (containing 70% starch). Corn can be processed by wet and dry milling. Wet milling produces starches, sweeteners (such as high fructose corn syrup), gluten feed, meal, dextrose, ethanol, and maltodextrins. Dry milling includes three processes, the stone grinding process, the dry-grind ethanol system, and the tempering degermination system. Stone grinding produces whole corn meal that is further processed to remove the oil. The dry-grind ethanol process produces ethanol through the fermentation of processed kernels, and a dried solubles by-product that is an important component of livestock feed. The tempering degermination system produces foods such as flaking grits, coarse and fine grits, meal, flour, oil, and hominy feed.
Corn is also used for masa production, which is the starting material for tortillas, tortilla chips, taco shells, and corn chips. Masa is produced through the cooking of corn in alkali, followed by grinding to produce dough.
Corn grain and by-products of wet and dry milling are used as animal feed. The whole corn plant can also be used as feed for ruminant animals. Corn plants are harvested at an appropriate stage and fed to animals or stored as silage.
Monsanto analyzed the composition of forage and grain from transgenic corn event MON 89034 and a non-transgenic control to assess whether the transgenic corn differs from non-transgenic corn. The non-transgenic control has a genetic background similar to that of MON 89034 but does not contain the cry1A.105 and cry2Ab2 genes. Monsanto used a randomized complete block design and analyzed corn grain and forage grown at five field sites, with three replicates per block. Additionally, Monsanto analyzed the grain and forage from 15 different conventional corn hybrids produced in the same field trials with MON 89034 and the non-transgenic control, with three different hybrids planted at each of the five sites. Monsanto used data derived from those 15 conventional hybrids as references to generate a 99% tolerance interval for each analyzed component. Monsanto states that such data illustrate the natural variability in commercially grown corn varieties and that comparing the level of each component of the transgenic corn to the respective 99% tolerance interval can provide a perspective for biological relevance in the context of natural variability in corn. Monsanto also compared the compositional data obtained for MON 89034 to the data contained in the International Life Science Institute (ILSI)2 Crop Composition Database and to values reported in the literature.
Monsanto applied a mixed model analysis of variance for its statistical analysis conducted on compositional data obtained for MON 89034 and the conventional comparator line. The statistical analysis was performed on analytical data from each individual field site as well as on data from all five field sites combined. Statistically significant differences were declared at the 5% significance level (p < 0.05). The level of each individual component of MON 89034 was compared to that of the non-transgenic control for each individual site and all sites combined, as well as to the 99% tolerance interval.
Monsanto determined the levels of the following components of forage from corn event MON 89034 and the non-transgenic control:
No statistically significant differences were found in the levels of all analyzed components between forage from MON 89034 and forage from the conventional control, with the exception of phosphorus. MON 89034 had statistically significantly higher levels of phosphorus than its control, but these levels were within the 99% tolerance intervals for the commercial varieties and within the ranges reported in the literature and the ILSI Crop Composition Database. Monsanto concluded that MON 89034 corn was not materially different than conventional corn varieties.
Monsanto measured the levels of the following components of mature grain3 from corn event MON 89034 and mature grain from the non-transgenic corn:
A list of specific components contained in each group is shown in Table 2
| Proximates* & Fiber | Minerals | Amino Acids | Fatty Acids | Anti-Nutrients | Secondary Metabolites | Vitamins |
|---|---|---|---|---|---|---|
|
ash* fat* protein* moisture* carbohydrates acid detergent fiber (ADF) neutral detergent fiber (NDF) total dietary fiber (TDF) |
calcium copper iron magnesium manganese phosphorus potassium zinc |
methionine cystine lysine tryptophan threonine isoleucine histidine valine leucine arginine phenylalanine glycine alanine aspartic acid glutamic acid proline serine tyrosine |
palmitic (16:0) palmitoleic (16:1) stearic (18:0) oleic (18:1) linoleic (18:2) linolenic (18:3) arachidic (20:0) eicosenoic (20:1) behenic (22:0) |
phytic acid |
p-coumaric acid ferulic acid |
vitamin B1 vitamin B2 folic acid (B9) niacin (B3) vitamin E vitamin B6 |
Monsanto conducted a statistical analysis of the analytical results combined from all field trials and found no statistically significant differences between MON 89034 and the conventional non-transgenic control, except for stearic acid and arachidic acid (calculated as a percentage of total fatty acids). However, differences in the levels of these fatty acids were small. The levels of all measured components were within the ranges reported in the literature and the ILSI Crop Composition Database and within the 99% tolerance interval. Monsanto found several statistically significant differences in the level of nutrients within the individual locations, but these differences were not consistent across all locations. Monsanto concluded that these differences were not biologically significant.
Monsanto has concluded that corn event MON 89034 is not materially different in composition, safety, wholesomeness, or any relevant parameter from corn now grown, marketed, and consumed. At this time, based on Monsanto’s data and information, the agency considers Monsanto’s consultation on corn event MON 89034 to be complete.
Karin Ricker, Ph.D.
(1)Cry1A.105 is a chimeric protein comprised of domains from Cry1Ab, Cry1Ac, and Cry1F from B.t. subsp. aizawai and the C-terminal portion from Cry1Ac B.t. subsp. kurstaki. The Cry2Ab2 protein is derived from B.t. subsp. kurstaki.
(2)Monsanto used version 3.0 (accessed on 8-27-06) of the ILSI Crop Composition Database in its analysis. The database is maintained by ILSI and can be accessed at http://www.cropcomposition.org.
(3)Monsanto analyzed forage and grain samples for a total of 77 components; 68 components in grain and nine components in forage. The nine components measured in forage corresponded to components that were also measured in grain. The levels of 16 components in grain were below the limits of quantification for the respective assays. These components are not considered key nutrients. Therefore, data for these components were not statistically analyzed and were not provided in the submission. The remaining 52 components that were above the limits of quantification are shown in Table 2.
