Promega AMV Reverse Transcriptase

Avian Myeloblastosis Virus Reverse Transcriptase (AMV RT) is a high-performance enzyme used in molecular biology applications to catalyze the synthesis of complementary DNA (cDNA) from an RNA template. AMV RT is especially useful for high-temperature reactions and is preferred for templates with high secondary structure, such as GC-rich RNA sequences. This enzyme provides stability and efficiency, enabling reliable synthesis of cDNA even at higher reaction temperatures (37–58°C).

AMV Reverse Transcriptase has RNase H activity, which degrades the RNA strand of RNA:DNA hybrids during first-strand cDNA synthesis, allowing for efficient synthesis of complementary DNA.

Key Features & Benefits:

1. High Concentration:
   • Cat.# M9004 contains 600 units of AMV RT at a concentration of 20–25 U/μL, providing high enzyme activity for efficient cDNA synthesis, even with difficult templates.
2. Temperature Stability:
   • AMV RT is well-suited for templates with high secondary structure (e.g., GC-rich regions). Its stability at higher temperatures (from 37°C to 58°C) makes it the preferred reverse transcriptase for challenging RNA templates that require elevated reaction temperatures to maintain activity and efficiency.
3. 5X Reaction Buffer:
   • The enzyme is supplied with a 5X Reaction Buffer, containing 250 mM Tris-HCl (pH 8.3), 250 mM KCl, 50 mM MgCl₂, 2.5 mM spermidine, and 50 mM DTT, which is optimized for efficient cDNA synthesis and primer extension.
4. RNase H Activity:
   • The RNase H activity of AMV RT enables it to degrade the RNA strand in RNA:DNA hybrids, ensuring the efficient synthesis of second-strand cDNA during the cDNA synthesis process.
5. Versatility:
   • This enzyme is suitable for a variety of applications, including RT-PCR, primer extension, and RNA sequencing.
6. Application Flexibility:
   • AMV RT is versatile for first- and second-strand cDNA synthesis, and can also be used in combination with Taq DNA polymerase for RT-PCR. Up to 10 μL of an RT reaction containing AMV RT and the supplied reaction buffer can be added to a 50 μL PCR reaction for amplification, enhancing workflow efficiency.

Applications:

1. First- and Second-Strand cDNA Synthesis:
   • AMV RT is widely used for first-strand cDNA synthesis in RT-PCR and RNA sequencing applications, as well as for second-strand cDNA synthesis.
2. RT-PCR:
   • Used for generating complementary DNA (cDNA) from RNA samples prior to PCR amplification, AMV RT facilitates the detection and analysis of RNA targets.
3. Primer Extensions:
   • Ideal for primer extension experiments, where the enzyme is used to extend primers for various genetic analyses.
4. RNA Sequencing:
   • Useful for preparing RNA samples for RNA sequencing, allowing the identification of gene expression profiles.
5. cDNA Library Construction:
   • Used in cDNA library construction for gene expression studies, functional genomics, and transcriptomics research.

Specifications:

Storage:

• Store at -20°C for long-term storage. The enzyme should be stored in aliquots to avoid repeated freeze-thaw cycles.

Promega’s AMV Reverse Transcriptase is an ideal choice for high-temperature cDNA synthesis and RT-PCR applications. Its stability at higher temperatures and efficient performance in difficult templates make it essential for cloning, gene expression analysis, and RNA sequencing. The enzyme’s RNase H activity also allows for efficient second-strand synthesis, making it suitable for a broad range of molecular biology applications.