This is an instruction manual for AccuScript High Fidelity 1st Strand cDNA Synthesis Kit
The Agilent AccuScript High Fidelity 1st Strand cDNA Synthesis Kit is designed to produce robust first-strand cDNA with superior accuracy and yield. It utilizes a Moloney murine leukemia virus (MMLV) reverse transcriptase derivative combined with a proofreading exonuclease, offering greater than three-fold higher accuracy compared to leading reverse transcriptases. The kit provides enough reagents for 50 reactions and includes essential components such as AccuScript High Fidelity RT, buffer, primers, dNTPs, DTT, and RNase Block inhibitor.
| Reactions per kit | 50 reactions |
|---|---|
| Buffer concentration | 10x |
| Oligo(dT) primer concentration | 0.5 ug/ul |
| Random primers concentration | 0.1 pg/wl |
| dNTP Mix concentration | 25 mM each dNTP |
| DTT concentration | 100 mM |
| RNase Block activity | 40 U/ul |
| Optimal RNA OD ratio | 1.8-2.0 |
| Recommended incubation time | 60 minutes |
| Inactivation temperature | 70°C |
RNA samples with an OD 60/289 of 1.8-2.0 are considered optimal.
A 60-minute incubation is recommended for most targets, though shorter times may suffice for some applications.
The reaction should be terminated by incubating at 70°C for 15 minutes to inactivate the enzyme.
It is recommended, but if omitted, the volume of water in the reaction must be increased accordingly.