M-MLV (H-) Reverse Transcriptase,UA BIOSCIENCE_specification/price/image

Product Detail
Company Profile

Product Specification

Synonyms	M-MLV[H-] RT，M-MLV[H-] Reverse Transcriptase
Expression System	E.coli
Molecular Weight	76 kDa (Reducing)
Purity	＞95% by SDS-PAGE
Conjugation	Unconjugated
Tag	His Tag
Storage Buffer	20 mM Tris-HCl、100 mM NaCl、1 mM DTT、0.1 mM EDTA、0.01% Nonidet® P-40、50% Glycerol pH 7.5 @ 25°C
Stability & Storage	Store at -25 ~ -15℃ for 2 years
Reference	[1] Blain, S. W. , and S. P. Goff . "Differential Effects of Moloney Murine Leukemia Virus Reverse Transcriptase Mutations on RNase H Activity in Mg and Mn." Journal of Biological Chemistry 271.3(1996):1448-54. [2] Narukawa, Yutaro , et al. "Improvement of Moloney murine leukemia virus reverse transcriptase thermostability by introducing a disulfide bridge in the ribonuclease H region." Protein Engineering, Design and Selection (2021).

Background

M-MLV (H-) Reverse Transcriptase is an RNA-dependent DNA polymerase with reduced RNase H activity. This enzyme can use RNA (when synthesizing cDNA) or single-stranded DNA as a template and initiate the synthesis of a complementary DNA strand from a primer. M-MLV(H-) reverse transcriptase has no 3´ → 5´ exonuclease activity. It can be used to synthesize first strand cDNA more efficient than the wild type M-MuLV。

Components

Storage Solution : 100 U/ul M-MLV(H-) Reverse Transcriptase、20 mM Tris-HCl、100 mM NaCl、1 mM DTT、0.1 mM EDTA、0.01% Nonidet® P-40、50% Glycerol pH 7.5 @ 25°C
5*Reaction Buffer: 250 mM Tris-HCl、375 mM KCl、15 mM MgCl2、50 mM DTT (pH 8.3 at 25°C)

Protocol

1. Genomic DNA was removed from the extracted cell total RNA by DNase I.
2. After incubating at 37 °C for 30 minutes, add 1µl 0.5 M EDTA (to the final concentration of 5mm) and inactivate at 75°C for 10 minutes.
3. Mix RNA sample, Random Primer and 1 ul M-MLV(H-) Reverse Transcriptase in a sterile RNase-free microfuge tube.
4. Incubate the 20 μl cDNA synthesis reaction was incubated at 25℃ for 5 minutes and then at 42℃ for 1 hour.
5. Inactivate the enzyme at 65°C for 20 minutes. The cDNA product can be directly fed into the qPCR reaction or stored at -20°C.

Guidelines

Please avoid repeated freeze-thaw cycles.

Unit Definition

One unit is defined as the amount of enzyme that will incorporate 1 nmol of dTTP into acid-insoluble material in a total reaction volume of 50 μl in 10 minutes at 37°C using poly(rA)•oligo(dT)18 as template.

AntBio is a biotechnology group company dedicated to serving life sciences, aiming to help scientists accelerate research and improve work efficiency. AntBio provides comprehensive and high-quality reagent tools for basic research, drug development, and diagnosis, including research grade antibodies, proteins, biochemical reagents, and assay kits. These research tools are widely used in different segments of life science research. The group company currently consists of three brands, Absin, Starter-Bio and UA-Bio.