Phosphomannose isomerase: An efficient selectable marker for plant transformation

Abstract

Phosphomannose isomerase (PMI) catalyzes the reversible interconversion of mannose 6-phosphate and fructose 6-phosphate. Plant cells lacking this enzyme are incapable of surviving on synthetic medium containing mannose as a carbon source. Maize, wheat and barley plants, genetically modified to express the Escherichia coli manA gene (pmi) under the control of a plant promoter, were able to survive selection on mannose-containing medium. Transformation frequencies averaged 45% for maize transformation via Biolisticse, 35% for maize Agrobacterium-mediated transformation, 20% for wheat, 3% for barley, and 2% for watermelon transformation. Moreover, the frequencies exceeded those obtained for maize and wheat using the pat or bar gene with Basta w selection. A preliminary safety assessment has been conducted for PMI. Purified PMI protein demonstrates no adverse effects in an acute mouse toxicity test. Purified PMI protein was readily digested in simulated mammalian gastric and intestinal fluids. Plants derived from sugar beet and corn cells that had been genetically modified to express the E. coli manA gene were evaluated for biochemical changes in mannose-associated pathways. No detectable changes in glycoprotein profiles were detected in PMI-transformed plants as compared to nontransgenic controls. The yield and nutritional composition of grain from PMI-transformed corn plants compared to their non-transformed isogenic counterparts were also determined and no statistically significant differences were found. The inherent safety of a system based on simple sugar metabolism coupled with high transformation frequencies for monocots make pmi an ideal selectable marker for plant transformation.

Expression of The Human Acidic Fibroblast Growth Factor in Transgenic Tomato and Safety Assessment of Transgenic Lines

“…Therefore, these two proteins bear no similarity to known allergenic proteins and the preliminary but not conclusive statement is that they do not represent a risk for allergenicity. Our results are similar to the results reported for PMI by Reed et al (54) and are an endorsement for the wide use of the gene for PMI as a selectable marker in plant genetic transformation.…”

Recent Advances in Fruit Species Transformation

“…Helena and Canino required the lower combination of mannose with sucrose (1,25 g/L mannose and 20 g/L sucrose) in comparison with other woody fruit trees to obtain the most effective selection procedure. Moreover, safety assessments were revealed that there is no any adverse effect of the enzyme on mammalian allergenicity and toxicity (Reed et al, 2001). …”

Application (Reference EFSA-GMO-UK-2005-11) for the placing on the market of insect-resistant genetically modified maize MIR604 event, for food and feed uses, import and processing under Regulation (EC) No 1829/2003 from Syngenta Seeds S.A.S on behalf of

“…PMI enzymes occur in a wide range of organisms including prokaryotes and eukaryotes such as bacteria, yeasts, animals, and humans, as well as plants, in which PMI is involved, for example, in glycoprotein synthesis. In MIR604 maize plants expressing PMI, no change in glycoprotein profiles has been observed (see section 4.1.2), indicating no effects of the introduction of PMI on the host plants' protein glycosylation (Reed et al, 2001). At the EFSA GMO Panel's request, the applicant also provided data on the pH-activity profile of PMI-0105, the bacterially produced analogue of the newly expressed PMI enzyme in maize MIR604.…”