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6% SDS-PAGE and stained with Coomassie blue.
To further examine the compatibility of the mirVana
PARIS Kit with tissues, the enrichment procedure was performed with mouse
brain or kidney that were stored for several months in RNAlater,
Ambion's tissue collection/stabilization solution. Previous studies showed
that both high quality protein and RNA are recovered from RNAlater-treated
samples [2, 6]. Analysis by denaturing acrylamide gel and Northern blot
of the RNA samples purified with the mirVana PARIS Kit confirm
that the procedure efficiently fractionates long and short RNA species
(Figure 6). mRNAs and 5.8S rRNA were detected only in the "depleted" fraction,
and the miRNAs let-7 or miR-124 were seen only in the "enriched" fraction.
Figure 6. Small RNA Enrichment from Mouse Tissues. Total RNA or fractions depleted or enriched in small RNA were isolated from RNAlater-treated mouse brain or kidney (~50 mg) with the mirVana PARIS Kit (A) 1 g of each fraction was resolved on a 15% denaturing acrylamide gel and stained with ethidium bromide. (B) mRNA, rRNA and miRNA expression was analyzed as described in Figure 4. This figure demonstrates the broad range of RNAs that can be obtained with this procedure, from miRNAs (~22 nt) to much larger mRNA species (in the kilobase range).
Versatile Isolation System
In summary, these results show
that high quality total RNA and protein are efficiently
recovered from cultured cells and several differ
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