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Development of a multiplex and semi-quantitative PCR assay for detection of genetically modified Roundup Ready Soybean in some representative food
基因改造生物（Genetically Modified Organisms, GMO）於近年來備受關注，目前已有不少基因改造作物於市場上流通。我國進口之大豆中已逾50%為基因改造大豆，其中又以Roundup ReadyTM品種為主；行政院衛生署目前已研擬「基因改造食品安全性評估方法」，並同時公告「基因改造食品標示辦法」，形成我國基因改造食品之管理準則。
本研究為開發多套式聚合酶鏈鎖反應（Multiplex PCR）及半定量(Semi-quantitative) 鑑定方法，評估其應用於檢驗基因改造大豆及市售產品之可行性。實驗過程針對Roundup ReadyTM基因改造大豆之轉殖基因設計不同引子，分別為35SP-F/R (143 bp)、NOS-F/R (189 bp)、EPSPS-F/R (314 bp)及35SP-F/CTP-R (256 bp)，並利用LEC-F/R (175 bp)作為內生性基因之鑑定。結果顯示，利用上述引子35SP-F/CTP-R及LEC-F/R進行多套式PCR方法鑑定基因改造大豆及市售產品，可鑑別出基因大豆之檢體，且靈敏度可達0.1%，此結果可達到行政院衛生署規定之5%基因改造成分(GM soya)的標示基準。此外，為瞭解市售產品中之GM soya比例，本實驗亦將市售產品之PCR產物量與不同GM soya比例之DNA標準曲線作比對進行半定量分析，以推斷市售產品中之GM soya比例。本研究所使用之多套式PCR及半定量方法可容易檢出基因改造大豆之檢體，並可大幅降低檢測時間及成本。期藉由此檢驗方法之建立，能對基因改造食品之檢驗與標示具有相當大的助益。|
Abstract Genetically modified organisms (GMO) have raised much attention in recent years. Uptoday, great deals of genetically modified organisms are sold in the market. Over 50% of imported soybeans, mainly Roundup ReadyTM soybean, in Taiwan were genetically modified. Department of Health (DOH), The Executive Yuan, Republic of China has published “Safety assessment methodology for Genetically modified foods (GMF)” and “Labeling act for Genetically modified foods (GMF)”, which is the foundation of management of GMF in Taiwan. This studied was to develope the qualitative and semi-quantitative detection method for genetically modified soybean by using the multiplex PCR technique. The potential application of the developed methods in detection of GM soybeans raw material or products were also evaluated. In this stduy, various primers for detection of cloned genes in Roundup ReadyTM, namely, 35SP-F/R (143 bp), NOS-F/R (189 bp), EPSPS-F/R (314 bp) and 35SP-F/CTP-R (256 bp), were artificially synthezied. In addtion, primer LEC-F/R (175 bp) was used for detection of species specificity of soybeans. The results showed that the samples containing genetically modified soybeans were detected by multiplex PCR with primers 35SP-F/CTP-R and LEC-F/R. Semi-quantitative analysis of PCR products was developed by using Gel-Pro Analyzer and Sigma Plot softwere with standard curves of DNA that contained various ratios of GMO. This method was used to estimate the amount of GM soybeans in retail products that was made of soybeans. Our results showed that GM soybeans were not only easily detected by multiplex PCR and semi-quantitative method the detection time and cost could also be reduced significantly.
|Appears in Collections:||食品暨應用生物科技學系|
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