The MON80100 line of corn (Zea mays) was developed through a specific genetic modification to be ECB (Ostrinia nubilalis) resistant. The novel variety produces a truncated version of the insecticidal protein, CryIA(b) derived from Bacillus thuriengensis. Delta-endotoxins, such as the CryIA(b) protein expressed in MON80100 corn, act by selectively binding to specific receptors localized on the brush border midgut epithelium of susceptible insect species. Following binding, cation-specific pores are formed that disrupt midgut ion flow and thereby cause paralysis and death. CryIA(b) and related endotoxins are insecticidal only to lepidopteran or coleopteran insects and their specificity of action is directly attributable to the presence of specific receptors in the target insects. There are no receptors for delta-endotoxins of B. thuringiensis on the surface of mammalian intestinal cells, therefore, livestock animals and humans are not susceptible to these proteins. The modified corn line is protected from ECB damage and permits farmers to reduce pesticide applications in the cultivation of corn.<br>The MON80100 corn line was created through direct DNA transformation by microparticle bombardment of plant cells with DNA-coated gold particles. Two plasmids were used for transformation, one of which contained a copy of the cryIA(b) gene from B. thuriengensis subsp. kurstaki. Constitutive expression of the cryIA(b) gene was controlled by inclusion of sequences from the 35S promoter of cauliflower mosaic virus (CaMV), the HSP70 intron and the 3’-polyadenylation signal of the nopaline sythase (nos) gene from Agrobacterium tumefaciens. The second plasmid contained open reading frames encoding the 5-enolpyruvylshikimate-3-phosphate synthase (EPSPS) enzyme from the common soil bacterium, Agrobacterium sp. CP4, and glyphosate oxidoreductase (goxv247) from Achromobacter sp. strain LBAA. Constitutive expression of these genes in plant cells was under the control of the CaMV 35S promoter and nos terminator. Post-translational targeting of the CP4 EPSPS and glyphosate oxidoreductase enzymes to the chloroplast was accomplished by fusion of the 5’-terminal coding sequences with the chloroplast transit peptide DNA sequences from the Arabidopsis thaliana EPSPS and ribulose-1,5-bisphosphate carboxylase genes, respectively. In addition, both plasmids contained sequences encoding the enzyme neomycin phosphotransferase II (NPTII) from the Tn5 transposon of Escherichia coli, strain K12. Expression of the neo gene was regulated by a bacterial promoter and was used as a selectable trait to screen transformed E. coli for the presence of the plasmid vector. The expression of CP4 EPSPS and glyphosate oxidoreductase activity were used as selectable traits to screen transformed plants for the presence of the cryIA(b) gene. Apart from the sequences encoding Cry1A(b), CP4 EPSPS and glyphosate oxidoreductase, no other plant-translatable DNA sequences were introduced into the plant genome. Southern analysis of genomic DNA from 5 generations of backcrosses of the MON80100 line to the B73 parental line and 4 generations of crosses to an unrelated inbred line confirmed stable inheritance of the novel trait. <br>The expression levels of CryIA(b) protein, CP4 EPSPS and glyphosate oxidoreductase were determined in the leaves, kernels and pollen of transgenic corn. In leaves and kernels, the amounts of CryIA(b) and CP4 EPSPS were to be 1.3 or 0.57 mg/g leaf fresh weight and 1.85 or 4.11 mg/g kernel fresh weight, respectively. There were no detectable amounts of CryIA(b) protein in the pollen from transgenic MON80100 corn. Also, glyphosate oxidoreductase and NPTII were not detected in either leaves or kernels from MON80100 corn.