Development of an event-specific Real-time PCR detection method for the transgenic Bt rice line KMD1

被引:54
作者
Babekova, Ruzha [2 ]
Funk, Tristan [1 ]
Pecoraro, Sven [1 ]
Engel, Karl-Heinz [2 ]
Busch, Ulrich [1 ]
机构
[1] Bayer Landesamt Gesundheit & Lebensmittelsicherhe, D-85764 Oberschleissheim, Germany
[2] Tech Univ Munich, Lehrstuhl Allgemeine Lebensmitteltechnol, D-85350 Freising Weihenstephan, Germany
关键词
GMO; Transgenic rice; Kemingdao 1 (KMD1); Event-specific Real-time PCR; SiteFinding-PCR; Hybrid amplicon; POLYMERASE-CHAIN-REACTION; SYNTHETIC CRY1AB GENE; YELLOW STEM BORER; GENETICALLY-MODIFIED RICE; BACILLUS-THURINGIENSIS BERLINER; GM-RICE; ENHANCED RESISTANCE; FIELD-EVALUATION; INDICA RICE; CHILO-SUPPRESSALIS;
D O I
10.1007/s00217-008-0981-0
中图分类号
TS2 [食品工业];
学科分类号
0832 ;
摘要
The present study describes an event-specific quantitative Real-time PCR detection method for the transgenic Bt rice line Kemingdao 1 (KMD1). This rice line which is not approved in any country so far is likely to be approved in China in the near future. The developed primers amplify a DNA sequence spanning the integration site of the genetic construct in KMD1. DNA sequence information of this unknown site necessary for primer design was achieved using SiteFinding-PCR technique. The specificity of the detection method was shown by testing a number of different transgenic and conventional plant varieties (e.g. rice LL 601, LL 62, Bt 63). As alternative to genomic DNA, we developed double target hybrid amplicons as synthetic calibration standards in Real-time PCR. These amplicons contained both one copy of the KMD1 event-specific sequence and one copy of a sequence of the rice reference gene gos9. The limit of quantification (LOQ) of the method was tested to be 0.05%.
引用
收藏
页码:707 / 716
页数:10
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