Transgene-induced RNA interference: a strategy for overcoming gene redundancy in polyploids to generate loss-of-function mutations.
Gene redundancy in polyploid species complicates genetic analyses by making the generation of recessive, loss-of-function alleles impractical. We show that this problem can be circumvented using RNA interference (RNAi) to achieve dominant loss of function of targeted genes. Arabidopsis suecica is an allotetraploid (amphidiploid) hybrid of A. thaliana and A. arenosa. We demonstrate that A. suecica can be genetically transformed using the floral dip method for Agrobacterium-mediated transformation. Transgenes segregate as in a diploid, indicating that chromosome pairing occurs exclusively (or almost so) among homologs and not among homeologs. Expressing a double-stranded (ds) RNA corresponding to the A. thaliana gene, decrease in DNA methylation 1 (DDM1) caused the elimination of DDM1 mRNAs and the loss of methylation at both A. thaliana- and A. arenosa-derived centromere repeats. These results indicate that a single RNAi-inducing transgene can dominantly repress multiple orthologs.
Lawrence
Richard J
RJ
Biology Department, Washington University, 1 Brookings Drive, St Louis, MO 63130, USA.
Pikaard
Craig S
CS
D017360:Arabidopsis / Q000235:genetics* / Q000502:physiology; D029681:Arabidopsis Proteins / Q000235:genetics* / Q000378:metabolism; D001483:Base Sequence; D004268:DNA-Binding Proteins / Q000235:genetics* / Q000502:physiology; D008969:Molecular Sequence Data; D009154:Mutation; D030821:Plants, Genetically Modified; D011123:Polyploidy; D034622:RNA Interference; D012231:Rhizobium / Q000235:genetics; D012689:Sequence Homology, Nucleic Acid; D013045:Species Specificity; D014157:Transcription Factors / Q000235:genetics* / Q000502:physiology; D014170:Transformation, Genetic / Q000235:genetics; D019076:Transgenes / Q000235:genetics*
D029681:Arabidopsis Proteins; C118957:DDM1 protein, Arabidopsis; D004268:DNA-Binding Proteins; D014157:Transcription Factors