title: Levels of DNA methylation and transcript accumulation in leaves of transgenic maize varieties.
abstract: BACKGROUND
  Prior to their release in the environment, transgenic crops are examined for their health and environmental safety. In addition, transgene expression needs to be consistent in order to express the introduced trait (e.g. insecticidal and/or herbicide tolerance). Moreover, data on expression levels for GM events are usually required for approval, but these are rarely disclosed or they are considered insufficient. On the other hand, biosafety regulators do not consider epigenetic regulation (e.g. DNA methylation, ncRNAs and histone modifications), which are broadly known to affect gene expression, within their risk assessment analyses. Here we report the results of a DNA methylation (bisulfite sequencing) and transgene transcript accumulation (RT-qPCR) analysis of four Bt-expressing single transgenic maize hybrids, under different genetic backgrounds, and a stacked transgenic hybrid expressing both insecticidal and herbicide tolerance traits.
  
  
  RESULTS
  Our results showed differences in cytosine methylation levels in the FMV promoter andcry2Ab2transgene of the four Bt-expressing hybrid varieties. The comparison between single and stacked hybrids under the same genetic background showed differences in the 35S promoter sequence. The results of transgene transcript accumulation levels showed differences in bothcry1A.105andcry2Ab2transgenes among the four Bt-expressing hybrid varieties. The comparison between single and stacked hybrids showed difference for thecry2Ab2transgene only.
  
  
  CONCLUSIONS
  Overall, our results show differences in DNA methylation patterns in all varieties, as well as in transgene transcript accumulation levels. Although the detection of changes in DNA methylation and transgenic accumulation levels does not present a safety issue per se, it demonstrates the need for additional studies that focus on detecting possible safety implications of such changes.
authors.affiliation: Department of Crop Science, Federal University of Santa Catarina, Florianópolis, Santa Catarina Brazil ; GenØk - Centre for Biosafety, Tromsø, Norway ; Institute for Plant Genetics, Faculty of Natural Sciences, Leibniz University of Hannover, Hannover, Germany.
authors.forename: Vinicius
authors.identifier: 0000-0002-5741-8016
authors.initials: V
authors.lastname: Vilperte
authors.affiliation: GenØk - Centre for Biosafety, Tromsø, Norway.
authors.forename: Sarah Zanon
authors.initials: SZ
authors.lastname: Agapito-Tenfen
authors.affiliation: GenØk - Centre for Biosafety, Tromsø, Norway ; Unit for Environmental Science and Management, Potchefstroom Campus, North West University, Potchefstroom, South Africa.
authors.forename: Odd-Gunnar
authors.initials: OG
authors.lastname: Wikmark
authors.affiliation: Department of Crop Science, Federal University of Santa Catarina, Florianópolis, Santa Catarina Brazil.
authors.forename: Rubens Onofre
authors.initials: RO
authors.lastname: Nodari
keywords: DNA methylation; Epigenetics; Genetic stability; Genetically modified organism; Risk assessment; Stacked GMO; Zea mays