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Prepare siRNA and miRNA Probes in Just 1 Hour
mirVana™ miRNA Probe Construction Kit
• Prepare your
probe in less than 1 hour
• Ideal for
siRNA/miRNA studies
• Synthesize
high specific activity RNA probes for RPA, Northern,
Southern, or in situ
hybridization
• No cloning
required
With the mirVana miRNA Probe Construction
Kit, short (<100 nt) radiolabeled or nonisotopically labeled
RNA probes can be generated in less than 1 hour. These probes
are ideal for the investigation of a variety of small RNAs. Radiolabeled
probes prepared with the mirVana miRNA
Probe Construction Kit have been successfully used for the detection
of small nuclear RNA (snRNA), small interfering RNA (siRNA),
micro RNA (miRNA), and mRNA by RPAs, Northerns and in situ hybridization
(see Detecting miRNA & siRNA). Nonisotopically labeled probes
generated with the kit have also been used to study the distribution
of
miRNA
or mRNA in tissues by in situ hybridization (See Analysis
of miRNA Distribution in Mouse Brain, right).
RNA Probe Synthesis Made Fast, Simple and Inexpensive
The mirVana miRNA Probe Construction
Kit includes reagents for preparation of in vitro transcription
templates as well as reagents for in vitro transcription.
For template preparation, all you need to supply is a short, inexpensive DNA
oligonucleotide specific for the target of interest that also includes an 8
base sequence complementary to the 3' end of the T7 promoter primer included
in the kit. This oligonucleotide is annealed to the provided T7 promoter primer,
and a 30 minute fill-in reaction with Klenow fragment generates the double-stranded
transcription template. The resulting template is simply mixed with the provided
T7 RNA Polymerase, rNTPs, and Transcription Buffer along with your choice of
labeled rNTP. The reaction is then incubated for as little as 10 minutes at
37°C to generate labeled probe.
Complete Kit for Probe Preparation
The kit includes enough reagents to prepare
30 dsDNA templates and to perform 30 in vitro transcription reactions.
Each template construction reaction
supplies sufficient template for up to 20 transcription reactions. The kit
also includes a functional positive control, DNase I to remove residual DNA
template, and Probe Elution Buffer to gel purify RNA transcripts.
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Figure 1. miR-16
and miR-22 Expression in Mouse Brain. In
situ hybridization was performed on coronal mouse forebrain
sections. In situ hybridized cells in the mouse brain
cortex (A,B,D,E; 20X magnification) and the head of
the caudate nucleus (C,F; 40X magnification) are indicated
by arrows. Nonisotopically labeled probes were prepared
with Ambion's MAXIscript™ Kit (1.5
kb probe) or with the mirVana miRNA
Probe Construction Kit (32 nt long probes, 22 nt target
specific sequence). |
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