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Integrated structure and event-specific real-time detection of transgenic cry1Ac/SCK rice Kefeng 6

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Abstract

Transgenic rice Kefeng 6 is a transformation event containing two insect-resistant genes, cry1Ac and SCK (modified CpTI gene) in China. In order to monitor the probable release of Kefeng 6 in the future and execute the labeling requirements, it is necessary to develop a rapid and reliable detection method. In this study, both the 5′ and 3′-junction sequences spanning the plant DNA and the integrated gene construct of the rice event Kefeng 6 were isolated by genome walking and long-distance PCR (LD-PCR), successively. Multiple copies of truncated SCK gene and cry1Ac gene were found to integrate into the host rice genome. The event-specific real-time detection method for Kefeng 6 event based on its 5′-junction sequence was established using one plasmid molecule pMD-KF6 containing both 5′-junction sequence and rice endogenous gene gos9 sequence as the reference material (RM) with an absolute limit of quantification (LOQa) around 10 template copies. Thereafter, three different transgenic amounts of w/w mixed samples (5, 1, and 0.5%, respectively) were quantified to assess the performance characteristics of the established real-time PCR method. The accuracy expressed as bias deviated from the 4.00–26.00%, the precision expressed as standard deviation (SD) and relative standard deviation (RSD) deviated from 0.03–0.19 and 3.42–4.76%, respectively. Based on the earlier results, we concluded that the qualitative and quantitative PCR assays were reliable and accurate for Kefeng 6 measurement, and the reference plasmid pMD-KF6 could be a good substitute for the reference material for Kefeng 6 quantification.

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Acknowledgments

This work was supported by the Fund of National Key Basic Research Developments Program of the Ministry of Science and Technology People’s Republic of China (2007CB109201) and the National Transgenic Plant Special Fund (2011ZX08011-001 and 2011ZX08012-002).

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Authors and Affiliations

  1. Inspection Test Center for Environmental Safety of Transgenic Crops (Beijing), Ministry of Agriculture, State Key Laboratory for Biology of Plant Diseases and Insect Pests Institute of Plant Protection, Chinese Academy of Agricultural Sciences, 100193, Beijing, China

    Changqing Su, Jiajian Xie, Xifeng Wang & Yufa Peng

  2. Department of Life Science, HengShui College, 053000, Hengshui, China

    Changqing Su

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  1. Changqing Su
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  2. Jiajian Xie
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  3. Xifeng Wang
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  4. Yufa Peng
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Correspondence to Jiajian Xie or Yufa Peng.

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Su, C., Xie, J., Wang, X. et al. Integrated structure and event-specific real-time detection of transgenic cry1Ac/SCK rice Kefeng 6. Eur Food Res Technol 232, 351–359 (2011). https://doi.org/10.1007/s00217-010-1399-z

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  • DOI: https://doi.org/10.1007/s00217-010-1399-z

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