title: Simultaneous knockdown of six non-family genes using a single synthetic RNAi fragment in Arabidopsis thaliana.
abstract: BACKGROUND
  Genetic engineering of plants that results in successful establishment of new biochemical or regulatory pathways requires stable introduction of one or more genes into the plant genome. It might also be necessary to down-regulate or turn off expression of endogenous genes in order to reduce activity of competing pathways. An established way to knockdown gene expression in plants is expressing a hairpin-RNAi construct, eventually leading to degradation of a specifically targeted mRNA. Knockdown of multiple genes that do not share homologous sequences is still challenging and involves either sophisticated cloning strategies to create vectors with different serial expression constructs or multiple transformation events that is often restricted by a lack of available transformation markers.
  
  
  RESULTS
  Synthetic RNAi fragments were assembled in yeast carrying homologous sequences to six or seven non-family genes and introduced into pAGRIKOLA. Transformation of Arabidopsis thaliana and subsequent expression analysis of targeted genes proved efficient knockdown of all target genes.
  
  
  CONCLUSIONS
  We present a simple and cost-effective method to create constructs to simultaneously knockdown multiple non-family genes or genes that do not share sequence homology. The presented method can be applied in plant and animal synthetic biology as well as traditional plant and animal genetic engineering.
authors.affiliation: Biosciences Division, Oak Ridge National Laboratory, Oak Ridge, TN 37831 USA ; KWS SAAT SE, Grimsehlstraße 31, 37555 Einbeck, Germany.
authors.forename: Olaf
authors.initials: O
authors.lastname: Czarnecki
authors.affiliation: Biosciences Division, Oak Ridge National Laboratory, Oak Ridge, TN 37831 USA.
authors.forename: Anthony C
authors.initials: AC
authors.lastname: Bryan
authors.affiliation: Biosciences Division, Oak Ridge National Laboratory, Oak Ridge, TN 37831 USA.
authors.forename: Sara S
authors.initials: SS
authors.lastname: Jawdy
authors.affiliation: Biosciences Division, Oak Ridge National Laboratory, Oak Ridge, TN 37831 USA.
authors.forename: Xiaohan
authors.initials: X
authors.lastname: Yang
authors.affiliation: Department of Plant Sciences, University of Tennessee, Knoxville, TN 37996 USA.
authors.forename: Zong-Ming
authors.initials: ZM
authors.lastname: Cheng
authors.affiliation: Biosciences Division, Oak Ridge National Laboratory, Oak Ridge, TN 37831 USA.
authors.forename: Jin-Gui
authors.initials: JG
authors.lastname: Chen
authors.affiliation: Biosciences Division, Oak Ridge National Laboratory, Oak Ridge, TN 37831 USA.
authors.forename: Gerald A
authors.initials: GA
authors.lastname: Tuskan