Gene Expression Analysis |
Transcriptome Discovery
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Transcriptome Analysis: ExactSTART™ Tools for Rapid and Efficient cDNA Preparation for Deep Sequencing, Cloning,
and Expression Profiling (940K PDF poster)
Applications
- Discover alternative transcription and polyadenylation sites of mRNAs.
- Unambiguously map the exact 5´ and 3´-ends of mRNAs.
- Identify and characterize splice variants and transcript homologs.
- Produce double-strand cDNA from eukaryotic mRNA for RACE or other RNA transcript
discovery method.
One common RACE template preparation procedure relies on the addition of non-template-coded
dNTPs to the 3´-end of the cDNA produced during reverse transcription
of the RNA. Such deoxynucleotide additions make it difficult to identify the
true 5´-end of the transcript. In addition, dNTPs are added to some cDNA
molecules more efficiently than others which may result in loss of transcripts
from the final population of RACE-amplified cDNA.
In contrast, the ExactSTART Eukaryotic mRNA 5´ & 3´-RACE Kit procedure uses
a rapid and improved "oligo-tagging" process to tag the exact 5´-nucleotide
of mRNAs and then produces 5´ and 3´-tagged cDNA that can be used
as is, converted to double-strand cDNA, or amplified by PCR to produce enriched
(amplified) ds cDNA for 5´ and 3´-RACE or other application such
as production of next-gen sequencing template.
The ExactSTART™ Eukaryotic mRNA 5´ & 3´ RACE Kit produces
5´ and 3´-end tagged cDNA or amplified double-strand cDNA (ds cDNA)
from 3´-polyadenylated RNAs (e.g., eukaryotic mRNA) that have:
| 5´-Cap / 3´-polyadenylated ends |
|
5´ m7GpppN-------[A]n 3´ or |
| 5´-monophosphorylated / 3´-polyadenylated ends |
|
5´ pN----------------[A]n 3´ or |
| 5´-triphosphorylated / 3´-polyadenylated ends |
|
5´ pppN-------------[A]n 3´ |
Benefits
- Accurate – Tag the exact 5´-nucleotide
of mRNAs to unambiguously map the true transcription site of
an mRNA. Unlike other 5´-RACE template production methods, the
ExactSTART approach does not add template-independent nucleotides during
the tagging process.
- Fast – Generate RACE template in less than a day.
- Easy – Simplified protocol requires no gel purification
steps.
- Flexible – Reactions generate cDNA, ds cDNA or amplified
ds cDNA for use as RACE template or other application such as production
of Next-gen sequencing template.
|
Figure 1 (click to enlarge). Schematic overview of the process for
the ExactSTART™ Eukaryotic mRNA 5´- & 3´-RACE
Kit. |
APex™ Heat-Labile Alkaline Phosphatase
ExactSTART™ End-Tagged Double-Strand cDNA Synthesis Kit for Small RNA
ExactSTART™ Full-Length cDNA Library Cloning Kit
ExactSTART™ Small RNA Cloning Kit
T4 RNA Ligase
Tobacco Acid Pyrophosphatase (TAP)
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