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The RT² miRNA First Strand Kit contains all reagents needed for converting
miRNA into first strand cDNA for real-time PCR. The simple one-step reaction not only performs both poly-(A) tailing and reverse transcription at
the same time. It also incorporates a proprietary and unique universal PCR
priming sequence providing a scalable analysis solution for multiple miRNA
sequences in the same cDNA preparation. The patent-pending miRNA First Strand
formulation improves the specificity of reverse transcription for mature miRNA
sequences over their precursors, primary transcripts, and mRNA targets. The kit
also provides greater control over RNA quality with its built-in External RNA
Control as well as enhanced sensitivity over other suppliers' miRNA first strand
cDNA synthesis kits. Due to the unique nature of the universal PCR priming
sequence, the RT² miRNA First Strand Kit is specifically designed for maximizing
the performance of the RT² miRNA PCR Array and Assay Systems.
Benefits of the RT² miRNA First Strand Kit
- Improved Specificity & Sensitivity: Proprietary reaction conditions do
not reverse transcribe miRNA precursors, primary transcripts, or mRNA - only
mature miRNA is detected
- Insure RT Efficiency & Consistency: Built-in External RNA Control
verifies lack of enzyme inhibitors and insures efficient reverse
transcription
- Easy and Fast: Everything included in one kit to complete first strand
cDNA synthesis in a one-step reaction
HOW IT WORKS:
1. A Poly-A Polymerase adds a poly-A tail to the miRNA sequence.
2. A primer, containing oligo(dT) and a universal PCR primer sequence, anneals
to the poly-adenylated RNA.
3. A Reverse Transcriptase synthesizes and appends to the primer DNA sequence
complementary to the miRNA.
4. The final first strand cDNA product contains the miRNA complement sequence, a
poly(dT) tract, and the universal PCR primer sequence. It is immediately ready
for a real-time PCR assay containing a miRNA-specific and the universal PCR
primer.
Control:
A built-in miRNA External RNA Control (ERC) helps test for inhibitors of
reverse transcription. Control elements on the RT² miRNA PCR Arrays
detect template synthesized from the ERC. A reproducible threshold cycle value
from this control across your samples indicates a consistently high level of RNA
quality and transcription efficiency giving you a greater degree of confidence
in your results.
Easy and Fast Sensitivity:
Combining poly(A) tailing and reverse transcription in one reaction and one
tube saves time and sample while reducing variability. The optimized reaction
conditions and buffer components maximize cDNA product yield using anywhere from
50 to 400 ng of input small RNA, and are most compatible with our RT² SYBR Green
qPCR Master Mixes. The universal PCR priming sequence incorporated during
reverse transcription also provides a scalable solution for analyzing the
expression of multiple miRNA sequences in the same cDNA preparation under
standardized conditions.
Complete RT² miRNA PCR System
Other sources of miRNA reverse transcription assay will not work with the
RT² miRNA PCR Arrays and Assays, and cDNA from the RT² miRNA First Strand Kit
cannot be used with other source of miRNA qPCR assays. Only the complete RT²
miRNA PCR System yields detectable real-time PCR results.
For more information, see our RT² miRNA PCR Array System User Manual
[here], or see the RT² miRNA First Strand Kit Specification Sheet
[here].
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