title: An efficient RNA interference screening strategy for gene functional analysis.
abstract: BACKGROUND
  RNA interference (RNAi) is commonly applied in genome-scale gene functional screens. However, a one-on-one RNAi analysis that targets each gene is cost-ineffective and laborious. Previous studies have indicated that siRNAs can also affect RNAs that are near-perfectly complementary, and this phenomenon has been termed an off-target effect. This phenomenon implies that it is possible to silence several genes simultaneously with a carefully designed siRNA.
  
  
  RESULTS
  We propose a strategy that is combined with a heuristic algorithm to design suitable siRNAs that can target multiple genes and a group testing method that would reduce the number of required RNAi experiments in a large-scale RNAi analysis. To verify the efficacy of our strategy, we used the Orchid expressed sequence tag data as a case study to screen the putative transcription factors that are involved in plant disease responses. According to our computation, 94 qualified siRNAs were sufficient to examine all of the predicated 229 transcription factors. In addition, among the 94 computer-designed siRNAs, an siRNA that targets both TF15 (a previously identified transcription factor that is involved in the plant disease-response pathway) and TF21 was introduced into orchids. The experimental results showed that this siRNA can simultaneously silence TF15 and TF21, and application of our strategy successfully confirmed that TF15 is involved in plant defense responses. Interestingly, our second-round analysis, which used an siRNA specific to TF21, indicated that TF21 is a previously unidentified transcription factor that is related to plant defense responses.
  
  
  CONCLUSIONS
  Our computational results showed that it is possible to screen all genes with fewer experiments than would be required for the traditional one-on-one RNAi screening. We also verified that our strategy is capable of identifying genes that are involved in a specific phenotype.
authors.affiliation: Department of Computer Science, National Tsing Hua University, Hsinchu, Taiwan.
authors.forename: Chih-Hung
authors.initials: CH
authors.lastname: Chang
authors.forename: Hsiang-Iu
authors.initials: HI
authors.lastname: Wang
authors.forename: Hsiang-Chia
authors.initials: HC
authors.lastname: Lu
authors.forename: Cheng-En
authors.initials: CE
authors.lastname: Chen
authors.forename: Hong-Hwa
authors.initials: HH
authors.lastname: Chen
authors.forename: Hsin-Hung
authors.initials: HH
authors.lastname: Yeh
authors.forename: Chuan Yi
authors.initials: CY
authors.lastname: Tang
mesh_terms: D016960:Agrobacterium tumefaciens / Q000235:genetics; D000465:Algorithms; D001483:Base Sequence; D019295:Computational Biology; D020224:Expressed Sequence Tags; D019668:Liliaceae / Q000235:genetics* / Q000276:immunology; D008969:Molecular Sequence Data; D010935:Plant Diseases / Q000235:genetics; D010940:Plant Proteins / Q000037:antagonists & inhibitors / Q000235:genetics* / Q000378:metabolism; D034622:RNA Interference; D034741:RNA, Small Interfering / Q000235:genetics*; D014157:Transcription Factors / Q000037:antagonists & inhibitors / Q000235:genetics* / Q000378:metabolism; D014170:Transformation, Genetic
chemical_list: D010940:Plant Proteins; D034741:RNA, Small Interfering; D014157:Transcription Factors