DNA interference: a simple and efficient gene-silencing system for high-throughput functional analysis in the fern adiantum.
RNA interference (RNAi) has become a powerful tool for determining gene function and is used in a wide variety of organisms. Since it is necessary to generate double-stranded RNA (dsRNA) as an inducer for RNAi, preparation of RNAi-inducing constructs is somewhat cumbersome and time consuming, especially for the thousands of genes used in a genome-wide analysis. To overcome these problems, we have developed a more convenient gene-silencing method in the fern Adiantum using double-stranded DNA (dsDNA) as a model system for functional analysis in plants. Delivery of dsDNA fragments homologous to an endogenous gene into gametophytic cells can induce sequence-specific gene silencing. As it only requires dsDNA fragments homologous to a target gene, PCR-amplified fragments are enough to trigger gene silencing. Maximum gene silencing efficiencies of >90% have been achieved for transformed plants. In addition, simultaneous transfer of dsDNA fragments corresponding to multiple genes still has a silencing effect for individual genes. We term this approach 'DNA interference'.
Kawai-Toyooka
Hiroko
H
Department of Biological Sciences, Graduate School of Science, Tokyo Metropolitan University, Hachioji, Tokyo, 192-0397 Japan.
Kuramoto
Chihiro
C
Orui
Kyoko
K
Motoyama
Kentaro
K
Kikuchi
Kazuhiro
K
Kanegae
Takeshi
T
Wada
Masamitsu
M
D035681:Adiantum / Q000235:genetics*; D004247:DNA / Q000235:genetics*; D015536:Down-Regulation; D018506:Gene Expression Regulation, Plant; D020868:Gene Silencing; D018390:Gene Targeting; D017343:Genes, Plant; D010641:Phenotype; D010940:Plant Proteins / Q000235:genetics / Q000378:metabolism; D010957:Plasmids; D015398:Signal Transduction
D010940:Plant Proteins; D004247:DNA