以聚合酵素鏈鎖反應（PCR）方法及市售免疫套組檢測四種基因改造玉米   全文下載

Detection of Four Types of Genetically Modified Maize by Polymerase Chain Reaction and Immuno-Kit Methods

本研究以PCR方法及市售免疫套組探討鑑別檢測四種基因改造玉米。針對Event 176 （Novartis 公司）、Bt11 （Novartis 公司）、MON810 （Monsanto公司）及Liberty （AgrEvo 公司）四種GM-玉米之殖入基因分別選定具產品特異性引子，進行PCR 方法檢測。用以鑑別GM-玉米之產品特異性引子共四對，分別為CDPK-cry（Event176 GM-玉米）、IV01-cry （Bt11 GM-玉米）、HS01-cry （MON810 GM-玉米）及CM03-PA01 （Liberty GM-玉米）。結果顯示，GM-玉米參考品以HS01-cry 引子檢測時，其最低檢測量為0.01% （w／w）， CDPK-cry 及IV01-cry 引子均為0.1% （w／w），CM03-PA01引子則為1% （w／w）。GM-玉米參考品分別再以市售免疫套組測試，Event176 、Bt11 及MON810三種GM-玉米皆呈現陽性反應。為瞭解食品玉米原料含GM-玉米之狀況，首先以市售免疫套組方法測試食品玉米原料等檢體共二十件，結果發現供作爆米花之玉米粒檢體十四件皆呈陰性反應，玉米碎檢體六件則為陽性反應；進一步再以PCR方法測試該二十件檢體，發現供作爆米花之玉米粒檢體可檢出二種GM-玉米品種，而玉米碎同件檢體，則可檢出四種GM-玉米品種或三種GM-玉米品種。綜合上述，本報告所使用之PCR方法能區分一般與四種基因改造玉米，市售免疫套組僅能供作檢體篩選用途，套組呈陽性或陰性反應之檢體，皆建議再配合PCR方法以確認與鑑別GM-玉米品種。同時，本研究得知玉米原料含有混合之不同品種GM-玉米。

To detect and identify four types of genetically modified (GM) maize, a polymerase chain reaction (PCR) assay and an immuno-assay kit were performed in this study. Primers specific to inserted genes in the Event 176 GM maize (Novartis company), Bt11 (Novartis company), MON810 (Monsanto company) and Liberty (AgrEvo company) were used to conduct the PCR assay. Four pairs of primers, namely, CDPK-cry (Event176), IV01-cry (Bt11), HS01-cry (MON810) and CM03-PA01 (Liberty) were used to identify the GM-maize. Results showed that the limit of detection for GM-maize were 0.01%, 0.1%, and 1% (w/w) as using HS01-cry, CDPK-cry and IV01-cry, and CM03-PA01 primers, respectively. The GM-maize references as well as 20 food-grade maize samples were also tested using a commercial immuno-kit. Three GM-maize references, Event176, Bt11 and MON810, and 6 corn-middling pellet samples were tested as positives; while 14 popcorn samples were negative. To confirm these results, the 20 food-grade maize samples were further tested by the PCR method. Two types of GM-maize were detected in popcorn samples; while 3~4 types of GM-maize were detected in the samples of corn middling pellets. The results of this study demonstrate that using PCR method is capable of differentiating four types of GM-maize from non-GM products; while the commercial immuno-kit can only be used for screening purpose. All samples, with positive or negative results as tested by immuno-kit, need to be further confirmed by the PCR method. The results of this study also reveal that marketed maize products are usually mixed with different types of GM-maize. 本研究以PCR方法及市售免疫套組探討鑑別檢測四種基因改造玉米。針對Event 176 （Novartis 公司）、Bt11 （Novartis 公司）、MON810 （Monsanto公司）及Liberty （AgrEvo 公司）四種GM-玉米之殖入基因分別選定具產品特異性引子，進行PCR 方法檢測。用以鑑別GM-玉米之產品特異性引子共四對，分別為CDPK-cry（Event176 GM-玉米）、IV01-cry （Bt11 GM-玉米）、HS01-cry （MON810 GM-玉米）及CM03-PA01 （Liberty GM-玉米）。結果顯示，GM-玉米參考品以HS01-cry 引子檢測時，其最低檢測量為0.01% （w／w）， CDPK-cry 及IV01-cry 引子均為0.1% （w／w），CM03-PA01引子則為1% （w／w）。GM-玉米參考品分別再以市售免疫套組測試，Event176 、Bt11 及MON810三種GM-玉米皆呈現陽性反應。為瞭解食品玉米原料含GM-玉米之狀況，首先以市售免疫套組方法測試食品玉米原料等檢體共二十件，結果發現供作爆米花之玉米粒檢體十四件皆呈陰性反應，玉米碎檢體六件則為陽性反應；進一步再以PCR方法測試該二十件檢體，發現供作爆米花之玉米粒檢體可檢出二種GM-玉米品種，而玉米碎同件檢體，則可檢出四種GM-玉米品種或三種GM-玉米品種。綜合上述，本報告所使用之PCR方法能區分一般與四種基因改造玉米，市售免疫套組僅能供作檢體篩選用途，套組呈陽性或陰性反應之檢體，皆建議再配合PCR方法以確認與鑑別GM-玉米品種。同時，本研究得知玉米原料含有混合之不同品種GM-玉米。