PCR Enzymes and Solutions
M-MLV Retro Transcriptase
Moloney Murine Leukemia Virus Reverse Transcriptase (M-MLV RT) is a RNA-dependent DNA polymerase used to generate first-strand complementary DNA (cDNA) from poly-mRNA, total RNA or viral RNA for use in downstream applications such as RT-(q)PCR or cDNA cloning. This enzyme contains several point mutations in the RNase H domain that enhances its thermostability and increases the cDNA yield of full-length transcripts (>5 kb) than wild-type M-MLV RT.
Catalog
Applications
- Preparation of cDNA libraries
- First-strand cDNA synthesis
- Quantitative real-time PCR (RT-qPCR)
Due to the purpose of this product, it is always RNase-free.
*For research use only.
Suggested reaction conditions
Recommended use of 200 units per 20 µL reaction. Denature RNA and primers by incubating at 65-70 °C for 5 min and mix them with dNTPs, MgCl2, provided buffer (Ref.; BMVRTHC) and M-MLV Reverse Transcriptase. Incubate the mixture at 45 °C for 30 min, inactivate the enzyme at 95 °C for 2 min and proceed with the generated cDNA.
Activity unit definition
One unit incorporates 1 nmol of dTTP into acid-precipitable material in 10 minutes at 37°C using poly(A)•oligo(dT)25 as template-primer.
Storage conditions
Once received, store the lyophilized product at Room Temperature, while buffers and MgCl2 should be stored at 4 °C or -20 °C. Once reconstituted, store M-MLV Reverse Transcriptase at -20 °C or -80 °C. Avoid multiple freeze/thaw cycles by storing multiple aliquots.
Buffer composition
This lyophilized product is provided with 1X M-MLV Reverse Transcriptase Reconstitution Buffer (ref.: RB-L3MVRTHC), 10X M-MLV Reverse Transcriptase Reaction Buffer (Ref.: BMVRTHC) and 100 mM MgCl2 (Ref.: MGCL2).
The lyophilized M-MLV Reverse Transcriptase contains 50,000 U. To obtain a final concentration of 200 U/µL, add 250 µL of reconstitution buffer (ref.: RB-L3MVRTHC) and gently pipette up and down. Aliquot into smaller volumes to avoid multiple freeze/thaw cycles.
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