ThermoScript™ RT-PCR System & Platinum® Taq DNA Polymerase

Catalog number: 11146032

Invitrogen™  Related applications: Reverse Transcription

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Description

ThermoScript™ Reverse Transcriptase (RT) is an avian RT with reduced RNase H activity designed for RT-PCR of the most highly structured, difficult RNA templates. The ThermoScript™ RT-PCR Systems are designed to copy RNA with secondary structure and to improve specificity of priming. In conjunction with PCR, the system can be used to detect the presence of rare messages or to quantitate the amount of specific mRNA from small numbers of cells. The systems contain ThermoScript™ RT, a cloned avian reverse transcriptase that has been engineered to have reduced RNase H activity and to have high thermal stability (up to 70°C). Each system is optimized for generating first-strand cDNA from total or poly(A)+ RNA (1) (Figure 1). ThermoScript™ RT-PCR Systems:

• Are optimized for RT-PCR
• Can be used with highly structured RNA or gene-specific primers
• Reduce RNase H activity for greater first-strand cDNA yield
• Amplify first-strand cDNA products directly without organic extractions or precipitations
• Include oligo(dT) and random hexamers to prime cDNA synthesis for flexible use
• Offer a choice of Platinum® Taq DNA Polymerase for higher specificity (2) (Figure 2) when amplifying templates up to 3 kb, or Platinum® Taq DNA Polymerase High Fidelity for amplifying templates up to 12 kb (Figure 3)
For Research Use Only. Not for use in diagnostic procedures.

Specifications

PCR Method: 2-Step RT-PCR
Reverse Transcriptase: ThermoScript™
Polymerase: Platinum® Taq DNA Polymerase
Hot Start: Built-In Hot Start
Sample Type (General): RNA
Product Size: 100 reactions
Detection Method: Gel Electrophoresis
Final Product Size(s): 3 kb or less
Primer-Probe Compatibility: Unlabeled Oligos

Contents & storage

• 60 µl Platinum® Taq DNA Polymerase
• 2 × 1.25 ml 10X PCR buffer (no magnesium)
• 1 ml 50 mM Magnesium Chloride
• 1.3 ml KB Extender
• 100 µl ThermoScript™ RT (15 U/µl)
• 500 µl 5X cDNA Synthesis Buffer
• 250 µl 0.1 M DTT
• 2 × 250 µl 10 mM dNTP Mix
• 100 µl RNaseOUT™ (40 U/µl)
• 100 µl Oligo (dT)20 (50 µM)
• 250 µl Random Hexamers (50 ng/µl)
• 1.25 ml DEPC-Treated Water
• 2 × 50 µl E. coli RNase H (2 U/µl)

Store at -20°C (stability can be extended by storage at -70°C).

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