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Double-Antibody Sandwich ELISA for the Detection of Transgenic Bar Gene Herbicide-Tolerant Soybean

LI Xiaoyu1, ZHANG Chunyu1, GUO Dongquan1, ZHANG Linlin2, YOU Qing3, DONG Yingshan1, WANG Yongzhi1,*, LI Qiyun1,*   

  1. 1. Jilin Key Laboratory of Agricultural Microbiology, Key Laboratory of Integrated Pest Management on Crops in Northeast,
    Institute of Plant Protection, Jilin Academy of Agricultural Sciences, Gongzhuling 136100, China;
    2. College of Plant Protection, Northeast Agricultural University, Harbin 150030, China;
    3. College of Agronomy, Jilin Agricultural University, Changchun 130118, China
  • Online:2016-02-25 Published:2016-02-23
  • Contact: WANG Yongzhi, LI Qiyun

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Abstract:

To rapidly and efficiently detect transgenic Bar herbicide-tolerant soybeans, we produced a monoclonal antibody
and a polyclonal antibody against phosphinothricin acetyltransferase (PAT) protein encoded by the Bar gene and then
utilized them to established a double-antibody sandwich ELISA system for the detection of PAT protein, which could
quantify PAT in different tissues and materials from transgenic Bar herbicide-tolerant soybeans. Reaction conditions were
optimized as follows: 0.125 μg/mL capture antibody was coated onto the microtiter plates at 4 ℃ overnight after incubation
at 37 ℃ for 1 h, the antigen was incubated at 37 ℃ for 1.5 h, and the detection antibody at 6.25 μg/mL was incubated
at 37 ℃ for 1.5 h. The detection sensitivity was 0.04 ng/mL for purified PAT and 8 ng/mL for crude soybean protein.
The coefficient of variation of reproducibility was less than 3%. This method has been successfully applied to detect the
expression of PAT in the root, stem, leaf, flower and seed of transgenic soybean

Key words: Bar, double-antibody sandwich ELISA, soybeans, detection

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