An oligonucleotide array to detect genetically modified events in potato

Abstract

For rapid and simultaneous detection of transgenic elements in genetically modified (GM) food crops, we explored DNA array technology. Forty-four oligonucleotide 23-to 31-mers were selected to use in an array on the basis of melting temperature and sequence specificity. Selected oligonucleotides consisted of DNA fragments corresponding to structural and regulatory elements and selectable markers used in developing transgenic crops, such as potato. Other oligonucleotides represented endogenous genes from potato to serve as positive controls and from heterologous crops, such as soybean and canola, to serve as negative controls. Amino-terminated oligonucleotides were hand-spotted on activated nylon membrane with a commercial spotting device. Target DNA was isolated from foliage of transgenic and nontransgenic crops, including potato, and labeled with digoxigenin-dUTP by random priming following restriction digestion to reduce DNA fragment size. Hybridization signals were visualized by an alkaline phosphatase anti-DIG-Fab conjugate and the chemiluminescent substrate, CDP-star. We detected the presence or absence of transgenic elements in transgenic and nontransgenic potato samples. Preliminary studies demonstrated that more specific and sensitive hybridization signals were generated from an oligonucleotide probe array than from a PCR product array. We anticipate that oligonucleotide probe arrays will be useful for regulatory monitoring of transgenic events.