title: Computational estimation and experimental verification of off-target silencing during posttranscriptional gene silencing in plants.
abstract: Successful application of posttranscriptional gene silencing (PTGS) for gene function study in both plants and animals depends on high target specificity and silencing efficiency. By computational analysis with genome and/or transcriptome sequences of 25 plant species, we predicted that about 50% to 70% of gene transcripts in plants have potential off-targets when used for PTGS that could obscure experimental results. We have developed a publicly available Web-based computational tool called siRNA Scan to identify potential off-targets during PTGS. Some of the potential off-targets obtained from this tool were tested by measuring the amount of off-target transcripts using quantitative reverse transcription-PCR. Up to 50% of the predicted off-target genes tested in plants were actually silenced when tested experimentally. Our results suggest that a high risk of off-target gene silencing exists during PTGS in plants. Our siRNA Scan tool is useful to design better constructs for PTGS by minimizing off-target gene silencing in both plants and animals.
authors.affiliation: Plant Biology Division, The Samuel Roberts Noble Foundation, Ardmore, Oklahoma 73401, USA.
authors.forename: Ping
authors.initials: P
authors.lastname: Xu
authors.forename: Yuanji
authors.initials: Y
authors.lastname: Zhang
authors.forename: Li
authors.initials: L
authors.lastname: Kang
authors.forename: Marilyn J
authors.initials: MJ
authors.lastname: Roossinck
authors.forename: Kirankumar S
authors.initials: KS
authors.lastname: Mysore
mesh_terms: D016960:Agrobacterium tumefaciens; D000818:Animals; D017360:Arabidopsis / Q000235:genetics* / Q000378:metabolism*; D018506:Gene Expression Regulation, Plant / Q000502:physiology*; D012275:Oryza / Q000235:genetics / Q000378:metabolism; D010940:Plant Proteins / Q000235:genetics / Q000378:metabolism*; D030821:Plants, Genetically Modified; D034622:RNA Interference; D015203:Reproducibility of Results; D012984:Software; D014026:Nicotiana / Q000235:genetics / Q000378:metabolism*
chemical_list: D010940:Plant Proteins