title: High-throughput amplification of mature microRNAs in uncharacterized animal models using polyadenylated RNA and stem-loop reverse transcription polymerase chain reaction.
abstract: This study makes a significant advancement on a microRNA amplification technique previously used for expression analysis and sequencing in animal models without annotated mature microRNA sequences. As research progresses into the post-genomic era of microRNA prediction and analysis, the need for a rapid and cost-effective method for microRNA amplification is critical to facilitate wide-scale analysis of microRNA expression. To facilitate this requirement, we have reoptimized the design of amplification primers and introduced a polyadenylation step to allow amplification of all mature microRNAs from a single RNA sample. Importantly, this method retains the ability to sequence reverse transcription polymerase chain reaction (RT-PCR) products, validating microRNA-specific amplification.
authors.affiliation: Institute of Biochemistry and Department of Biology, Carleton University, Ottawa, Ontario K1S 5B6, Canada.
authors.forename: Kyle K
authors.initials: KK
authors.lastname: Biggar
authors.affiliation: Institute of Biochemistry and Department of Biology, Carleton University, Ottawa, Ontario K1S 5B6, Canada.
authors.forename: Cheng-Wei
authors.initials: CW
authors.lastname: Wu
authors.affiliation: Institute of Biochemistry and Department of Biology, Carleton University, Ottawa, Ontario K1S 5B6, Canada. Electronic address: kenneth_storey@carleton.ca.
authors.forename: Kenneth B
authors.initials: KB
authors.lastname: Storey
mesh_terms: D002001:Adipose Tissue, Brown / Q000378:metabolism; D000818:Animals; D001483:Base Sequence; D006801:Humans; D055029:Inverted Repeat Sequences / Q000235:genetics*; D051379:Mice; D035683:MicroRNAs / Q000096:biosynthesis* / Q000235:genetics; D023421:Models, Animal; D012333:RNA, Messenger / Q000235:genetics*; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods*
chemical_list: D035683:MicroRNAs; D012333:RNA, Messenger
keywords: Hibernation; Ictidomys tridecemlineatus; MicroRNA amplification; Polyadenylation; RT–PCR