Principle
This product employs pre-mixing technology (pre-mixed with TRUEscript H_RTase / RNase Inhibitor Mix) using RNA as the template to synthesize the cDNA strand of circRNA efficiently. The point-mutated M-MuLV reverse transcriptase TRUEscript H_RTase with lost activity of RNase H, which is cloned through genetic recombination technology.
The wild-type M-MuLV contains RNase H activity, which can catalyze the degradation of RNA in the DNA/RNA hybrid. Therefore, during the synthesis reaction of the first strand of cDNA, it may degrade the template RNA in the RNA/DNA hybrid. The RNase H activity of (M-MuLV [RNase H_] ) is absent, it has stronger extension ability and stability compared with M-MuLV , so can be used for the synthesis of longer cDNAs.
The RNase R in this product can break down almost all linear RNA molecules, but it is not capable of digesting circular RNA, loop structures or double-stranded RNA molecules with a 3' protruding end shorter than 7 nucleotides. It can be used for gene expression and alternative splicing studies, and it can break down linear RNA to enrich circular RNA (circRNAs) or loop structure RNA (lariat RNA).
Specification
Components | 4205A (50T) | 4205B (100T) |
5× TRUE Reaction Mix | 200 μL | 400 μL |
RNase R | 250 U | 500 U |
10× Reaction Buffer | 1 mL | 2 mL |
Random primer (N6) | 50 μL | 100 μL |
RNase free H2O | 1 mL | 1.5 mL |
Application
It is widley used for enriching circRNAs and synthesizing cDNA strands.
Unit Deinition
Under the standard reaction system, the amount of enzyme required to convert 1 μg of poly(A) into acid-soluble nucleotides at 37℃ for 10 min is defined as one activity unit (U).
Feature
The length of the synthesized cDNA fragment can reach up to 12kb.