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New Tools for Small RNA Analysis

p://www.ambion.com/prod/fcrna">"miRNA certified" FirstChoice RNAs.


Sensitive Detection of Small RNAs
Although Northern analysis can be used to detect and quantitate small RNA, the technique is labor-intensive and, more importantly, not very sensitive. Ambion scientists have developed a sensitive assay that has been optimized for small RNA quantitation--the patented mirVana miRNA Detection Kit--which is based on solution hybridization and subsequent RNase protection. Short radiolabeled RNA probe is simply mixed with total RNA in an optimized hybridization buffer and all single-stranded RNAs (e.g. unhybridized probe, mRNAs, rRNA) are then digested by a mixture of RNase A and T1. Probe:target RNA hybrids are quickly recovered using a patented single-step RNase inactivation/precipitation procedure, run on a denaturing polyacrylamide gel, and visualized by autoradiography. In this assay, full-length and protected probes are easily distinguished by the presence of a short sequence at the 5' or 3' end of probes that is not complementary to the RNA target (Figure 2).

Figure 2. The mirVana miRNA Detection Kit for small RNA Detection


The mirVana miRNA Detection Kit has been used to detect miRNAs and siRNAs in a variety of tissues and cells (see poster for more information). The sensitivity and specificity of the assay is illustrated in Figure 3 where miR-124 miRNA was detected only in human brain total RNA. This experiment confirms the tissue-specific expression
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