title: Cloning and profiling of small RNAs from cucumber mosaic virus satellite RNA.
abstract: RNA silencing is not only a gene regulation mechanism that is conserved in a broad range of eukaryotes but also an adaptive immune response against foreign nucleic acids including viruses in plants. A major feature of RNA silencing is the production of small RNA (sRNA) of 21-24 nucleotides (nt) in length from double-stranded (ds) or hairpin-like (hp) RNA by Dicer-like (DCL) proteins. These sRNAs guide the binding and cleavage of cognate single-stranded (ss) RNA by an RNA silencing complex. Like all plant viruses and subviral agents, replication of viral satellite RNAs (satRNAs) is associated with the accumulation of 21-24 nt viral small interfering RNA (vsiRNA) derived from the whole region of a satRNA genome in both plus and minus-strand polarities. These satRNA-derived siRNAs (satsiRNAs) have recently been shown to play an important role in the trilateral interactions among host plants, helper viruses and satRNAs. Here, we describe the cloning and profile analysis of satsiRNAs from satRNAs of Cucumber mosaic virus (CMV). We also describe a method to minimize the strand bias that often occurs during vsiRNA cloning and sequencing.
authors.affiliation: State Key Laboratory of Plant Genomics and National Center for Plant Gene Research, Institute of Microbiology, Chinese Academy of Sciences, Beijing, 100101, China.
authors.forename: Yuan-Yuan
authors.initials: YY
authors.lastname: Fang
authors.forename: Neil A
authors.initials: NA
authors.lastname: Smith
authors.forename: Jian-Hua
authors.initials: JH
authors.lastname: Zhao
authors.forename: Joanne R M
authors.initials: JR
authors.lastname: Lee
authors.forename: Hui-Shan
authors.initials: HS
authors.lastname: Guo
authors.forename: Ming-Bo
authors.initials: MB
authors.lastname: Wang
mesh_terms: D002874:Chromosome Mapping / Q000379:methods; D003001:Cloning, Molecular; D017799:Cucumovirus / Q000235:genetics*; D004591:Electrophoresis, Polyacrylamide Gel / Q000379:methods; D015967:Gene Expression Regulation, Viral; D019244:RNA, Satellite / Q000302:isolation & purification; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods
chemical_list: D019244:RNA, Satellite