QUANTIFICATION OF TRANSGENIC PLANT MARKER GENE PERSISTENCE IN THE FIELD

Methods were developed to monitor persistence of genomic DNA in decayi ng plants in the field. As a model, we used recombinant neomycin phosp hotransferase II (rNPT-II) marker genes present in genetically enginee red plants. Polymerase chain reaction (PCR) primers were designed, com plementary to 20-bp sequences of the nopaline synthase promoter in a t ransgenic tobacco and the cauliflower mosaic virus 35S promoter in a t ransgenic potato. The PCR reverse primer was complementary to a 20-bp sequence of the N-terminal NPT-II coding region. The PCR protocol allo wed for quantification of as few as 10 rNPT-II genes per reaction. We analysed rNPT-II marker gene amounts in samples obtained from two fiel d experiments performed at different locations in Oregon. In transgeni c tobacco leaves, buried at 10 cm depth in a field plot in Corvallis, marker DNA amount dropped to 0.36% during the first 14 days and was de tectable for 77 days at a final level of 0.06% of the initial amount. Monitoring of residual potato plant litter, from the soil surface of a test field in Hermiston, was performed for 137 days. After 84 days ma rker gene amounts dropped to 2.74% (leaf and stem) and 0.50% (tuber) o f the initially detected amount. At the final sample date 1.98% (leaf and stem) and 0.19% (tuber) were detectable. These results represent t he first quantitative analysis of plant DNA stability under field cond itions and indicate that a proportion of the plant genomic DNA may per sist in the field for several months.