Description
The EZBioscience® microRNA Reverse Transcription Kit uses the method of Poly (A) tailing reaction and reverse transcription reaction in a single step assay to synthesize the first strand cDNA of miRNAs. Mature miRNAs from total RNA are modified by extending the 3' end of the mature transcript through Poly(A) addition using E. coli Poly (A) Polymerase. The modified miRNAs then undergo universal reverse transcription using Oligo (dT)-universal tag primer to synthesize the first strand cDNA for all miRNAs. The kit can be used with microRNA probe qPCR Kits (Catalog No.: EZB-miProbe/EZB-miProbe-R0/EZB-miProbe-R1/EZB- miProbe-R2).
The kit contains three tubes of reagents: gDNA Remover, miRNA RT Enzyme Mix, and 4× miRNA RT Buffer.
Among them, gDNA Remover mainly includes concentrated DNase and buffer. It can degrade more than 95% of residual genomic DNA only needing to react at room temperature (19 ~ 27°C) for 5 minutes, which greatly reduces the interference to the results.
The miRNA RT Enzyme Mix mainly contains E. coli Poly(A) Polymerase, reverse transcriptase, and RNase Inhibitor. E. coli Poly (A) Polymerase not only has efficient Poly (A) tailing reaction efficiency, but also specifically recognizes mature single-stranded miRNA, thereby avoiding reverse transcription reaction of miRNA precursors with double-stranded structure. The mutant M-MLV reverse transcriptase has strong anti-interference ability and amplification ability.
The 4× miRNA RT Buffer reagent contains all raw materials and primers for Poly(A) tailing reaction and reverse transcription reaction, including Oligo(dT)-universal tag primer, buffer and dNTPs, and has been carefully optimized to ensure Poly (A) tailing reaction and reverse transcription reaction efficiently.
Components
Components | EZB-miRT4-S (20 Rxns) | EZB-miRT4-L (50 Rxns) |
gDNA Remover | 22 μl | 55 μl |
miRNA RT Enzyme Mix | 44 μl | 110 μl |
4× miRNA RT Buffer | 110 μl | 275 μl |
Nuclease free ddH2O | 1 ml | 1 ml |
Storage
Store at -20°C.
Features
1. Fast and simple procedure: This kit uses the method of Poly (A) tailing reaction and reverse transcription reaction in a single step assay to synthesize the first strand cDNA of miRNAs. There is no need to design additional reverse transcription primers. high-quality cDNA containing all miRNAs can be obtained in 30 minutes.
2. High specificity: Poly(A) Polymerase not only has high A-tailing efficiency, but also specifically recognizes mature single-stranded miRNA, thus avoiding the reverse transcription reaction of miRNA precursors with double-stranded structure.
3. High efficiency: This kit uses an optimized combination of high-efficiency Poly(A) Polymerase and reverse transcriptase, which can efficiently carry out Poly (A) tailing and reverse transcription reactions. Compared with common commercial reverse transcription products, this kit reverses cDNA with the smallest quantitative CT value and has excellent reverse transcription efficiency.
4. The kit should be used with EZBioscience® EZ-Probe qPCR Master Mix for microRNA kits(Catalog No.: EZB-miProbe/EZB-miProbe-R0/EZB-miProbe-R1/EZB- miProbe-R2).
For detailed information, please check the product manual.
