PCR |
End-Point RT-PCR
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Applications
- High-sensititivity detection of RNA, including RNA present at low copy
number.
- RT-PCR applications where sensitivity is the primary concern.
- Multiplex RT-PCR.
The MasterAmp™ RT-PCR Kit for High Sensitivity is a single enzyme,
single-tube RT-PCR system (Fig. 1). It is capable of amplifying RNA from
as little as 1 pg of total cellular RNA (Fig. 2). It is ideal for amplifying
low-copy-number RNA transcripts.
The MasterAmp RT-PCR Kit for High Sensitivity uses RetroAmp™ RT DNA
Polymerase, a thermostable enzyme that has peak reverse transcriptase activity
at ≥60°C and lacks RNase H activity. Reverse transcription at higher
temperatures reduces RNA secondary structure and increases the specificity
and sensitivity of cDNA synthesis. The sensitivity of this kit is further
augmented because both RT and PCR occur in a single tube. All cDNA synthesized
during RT is used as a template for PCR amplification. |
Figure 1. Different RT-PCR protocols that can be used with the MasterAmp™ High
Fidelity RT-PCR Kit. |
The kit also incorporates MasterAmp PCR Enhancer with betaine,* which further
improves the specificity, consistency, and sensitivity of RT-PCR by reducing
pauses and stops during DNA synthesis.
Note: Although the MasterAmp RT-PCR Kit for High Sensitivity provides unsurpassed
RT-PCR sensitivity, the MasterAmp High Fidelity RT-PCR Kit should be used to
amplify RNA by RT-PCR for cloning, sequencing, or expression, when high fidelity
is most important.
Benefits
- High-sensitivity RT-PCR from as little as 1 pg of total cellular RNA (Fig.
2).
- Outperforms similar products from other vendors.
- Thermostable RetroAmp™ RT DNA Polymerase enables RT to be performed
at higher temperature, which minimizes RNA secondary structure.
- Use of one enzyme in a one-step and one-tube reaction with no sample transfers
reduces the chance for cross-contamination.
- The MasterAmp PCR Enhancer technology increases RT-PCR sensitivity and specificity.
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| Figure 2. RT-PCR amplification of human chorionic gonadotropinalpha
(hCGα) from decreasing amounts of placental total cellular RNA using
MasterAmp™ RT-PCR Kit for High Sensitivity. RNA was isolated using
the MasterPure™ Complete DNA and RNA Purification Kit. A 40-cycle amplification
was performed using decreasing amounts of total RNA following the protocol
supplied. Lane M, 100-bp ladder. |
Figure 3. Human CGα was amplified from various amounts
of total human placental RNA using MasterAmp™ RT-PCR Kit for High Sensitivity
or kits from other manufacturers. All experiments were carried out strictly
according to each manufacturer's protocols. |
*Covered by issued and/or pending patents.
MasterAmp™ RT-PCR Kit for High Sensitivity. Purchase of this product
includes an immunity from suit under patents specified in the product insert
to use only the amount purchased for the purchaser's own internal research.
No other patent rights (such as 5´ Nuclease Process patent rights) are
conveyed expressly, by implication, or by estoppel. Further information on
purchasing licenses may be obtained by contacting the Director of Licensing,
Applied Biosystems, 850 Lincoln Centre Drive, Foster City, California 94404,
USA.
2´-Deoxyribonucleoside-5´-Triphosphate Solutions
FailSafe™ GREEN and PROBES, TAQurate™ GREEN and PROBES, MasterAmp™ GREEN, TAQXpedite™ GREEN (discontinued)
FailSafe™ PCR System
MasterAmp™ High Fidelity RT-PCR Kit
MasterPure™ Complete DNA and RNA Purification Kit
MonsterScript™ 1st-Strand cDNA Synthesis Kit
MonsterScript™ Reverse Transcriptase
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