The agricultural biotechnology industry applies polymerase chain reaction (PCR) technology at numerous points in product development. Commodity and food companies as well as third-party diagnostic testing companies also rely on PCR technology for a number of purposes. The primary use of the technology is to verify the presence or absence of genetically modified (GM) material in a product or to quantify the amount of GM material present in a product. This article describes the fundamental elements of PCR analysis and its application to the testing of grains. The document highlights the many areas to which attention must be paid in order to produce reliable test results. These include sample preparation, method validation, choice of appropriate reference materials, and biological and instrumental sources of error. The article also discusses issues related to the analysis of different matrixes and the effect they may have on the accuracy of the PCR analytical results.

Immunoassays for biotechnology engineered proteins are used by AgBiotech companies at numerous points in product development and by feed and food suppliers for compliance and contractual purposes. Although AgBiotech companies use the technology during product development and seed production, other stakeholders from the food and feed supply chains, such as commodity, food, and feed companies, as well as third-party diagnostic testing companies, also rely on immunoassays for a number of purposes. The primary use of immunoassays is to verify the presence or absence of genetically modified (GM) material in a product or to quantify the amount of GM material present in a product. This article describes the fundamental elements of GM analysis using immunoassays and especially its application to the testing of grains. The 2 most commonly used formats are lateral flow devices (LFD) and plate-based enzyme-linked immunosorbent assays (ELISA). The main applications of both formats are discussed in general, and the benefits and drawbacks are discussed in detail. The document highlights the many areas to which attention must be paid in order to produce reliable test results. These include sample preparation, method validation, choice of appropriate reference materials, and biological and instrumental sources of error. The article also discusses issues related to the analysis of different matrixes and the effects they may have on the accuracy of the immunoassays.

Characterization of embryogenesis-related proteins in alfalfa (Medicago sativa). -Physiol, Plant, 96: 585-592.Large samples of embryogenic ceils can be obtained from tetraploid alfalfa (Medicago sativa L,) suspension cultures following induction by auxin and culture on growth regulator-free medium, A one-dimensional sodium dodecyl sulphate-polyacr}.'lamide gel electrophoresis (SDS-PAGE) comparison of the polypepiides synthesized by embryogenic and nonembryogenic alfalfa genotypes during inductive conditions identified a 50-kDa polypeptide band that was specific to embryogenic genotypes. Two-dimensional (iso electric focusing SDS-PAGE) electrophoretic separations resolved the 50-kDa baad into three 50-kDa polypeptides with different iso-electric points, A comparison of the polypeptides in the soluble and microsomal membrane fractions from embrj'ogenic cell masses of an embryogenic genotype suggested that the 50-kDa peptides are membrane associated. The in vitro translation products from RNAs of an embryogenic genotype had a 5Q-kDa band that was absent from the in vitro hanslation products obtained from RNAs of a nonembryogenic genotype. Therefore, we concluded thai the membrane-localized 50-kDa proteins are involved in somatic embryogenesis in alfalfa and that their expression is transcriptionally determined.

Many countries are developing or implementing regulatory requirements to monitor for the presence of genetically modified (GM) materials in seeds, grain, and derived food products using DNA and protein-based methods. There is no published report on the detection of different GM transgenes in canola, and this study is aimed at developing qualitative PCR methods for the three major GM transgenes commercially available in canola. Primer sequences were generated from GenBank and previously published information to develop a polymerase chain reaction (PCR) detection method for Roundup Ready (glyphosate tolerance, GT73 event), Liberty Link (glufosinate ammonium tolerance, HCN92 event), and BX (Bromoxynil tolerance, OXY235 event) canola varieties. On using PCR, two primer pairs for each of the GT73 and HCN92 and one primer pair for OXY235 amplified specific amplicons for the three GM transgenes. All three GM transgenes were detected simultaneously by multiplexing the five primer pairs in a single PCR reaction. Multiplexing of the five primer pairs for DNA prepared at 1% (one GM seed in 99 non-GM seeds) and 0.5% (one GM seed in 199 non-GM seeds) levels generated the expected DNA fragments for GT73, HCN92, and OXY235. This information will lay the groundwork for the development of a quantitative PCR assay for canola transgenic events.

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