- Prepare miRNA samples for microarray analysis
- Label each miRNA with multiple fluorescently-labeled molecules for enhanced sensitivity
- Part of Ambion's mirVana™ miRNA Array System
The mirVana miRNA Labeling Kit (patent pending) is designed for the preparation of microRNA (miRNA) samples for microarray analysis using a
simple and effective end-labeling strategy. In the first step of the mirVana miRNA Labeling Kit procedure,
E. coli Poly(A) Polymerase and a mixture of unmodified and amine-modified
nucleotides are used to add a 20–50 nucleotide tail to the 3' end of each
miRNA in the sample. The amine-modified miRNAs are then purified and coupled to amine-reactive labeled moieties such as NHS-ester
CyDye™ fluors (Amersham Biosciences) to produce a fluorescently-labeled sample for microarray analysis.
An important benefit of
the tailing strategy is that each miRNA obtains multiple fluorescently-labeled molecules. Since most miRNAs comprise less than 0.0001%
of a total RNA sample, this high degree of labeling is very important for
obtaining sensitive and accurate miRNA microarray data.
Note: Plant miRNAs cannot be labeled with this kit due to endogenous 3' methylation. For more information, see reference below.
miRNA samples
labeled using the mirVana miRNA Labeling Kit can be hybridized
to a mirVana™ miRNA Bioarray or to miRNA arrays prepared using the mirVana™ miRNA Probe Set. Together these products comprise the Ambion mirVana™ Array System, an efficient, user-friendly platform for miRNA expression profiling that results in sensitive, accurate, and highly reproducible data (see Figure 1). Start the process by isolating miRNA-containing total RNA from cells and tissues using the mirVana™ miRNA Isolation Kit and enriching for small RNA using the flashPAGE™ Fractionator and Reaction Clean-up Kit.
The mirVana miRNA Labeling Kit contains sufficient reagents for the labeling of 20 miRNA samples and includes a positive control. The reactive dye(s) (e.g. CyDye) is not included.
Reference
Yu B, Yang Z, Li J, Minakhina S, Yang M, Padgett RW, Steward R, Chen X. (2005)
Methylation as a crucial step in plant microRNA biogenesis.
Science. 307:932–5. [abstract]
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