Isolation and biochemical analysis of plant small RNAs.
Small RNAs, defined as noncoding 20-30-nt-long RNAs, are instrumental regulators of cellular processes in most eukaryotes. In this chapter we describe three methods for extracting small RNA from cells: a general method, one plant specific and a third particular to conifers. Further, protocols are given for the analysis and quantification of small RNAs using polyacrylamide gel-based approaches. A native streptavidin gel-shift assay, useful for measuring the relative amounts of multiple small RNAs simultaneously, is presented. To further characterize small RNAs biochemically, a sodium periodate assay probing for 2', 3' hydroxyl groups on the 3' terminus of small RNAs is outlined.
Institute of Molecular Systems Biology, Eidgenössische Technische Hochschule Zürich, Zürich, Switzerland.
H Alexander
HA
Ebhardt
Mariana Oviedo
MO
Ovando
Peter J
PJ
Unrau
D004591:Electrophoresis, Polyacrylamide Gel / Q000379:methods; D024202:Electrophoretic Mobility Shift Assay / Q000379:methods; D035683:MicroRNAs / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification; D010944:Plants / Q000235:genetics*; D018749:RNA, Plant / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification; D034741:RNA, Small Interfering / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification; D019809:Streptavidin
D035683:MicroRNAs; D018749:RNA, Plant; D034741:RNA, Small Interfering; D019809:Streptavidin