|Cat# BB-E0045||50 Reactions|
Super RT Easy Kit.PDF DOWNLOAD
Murine Leukemia Virus Reverse Transcriptase (MuLV RT) is an RNA-dependent DNA polymerase that can be used in cDNA synthesis with long messenger RNA templates (>5kb). The enzyme is isolated from E. coli expressing modified pol gene of MuLV on a plasmid.
This modified version of RT is thermotolerant.
♦ Super RTEasy Enzyme mix: 100 μl (Green Cap)
♦2x Reaction Mix (contains dNTPs, Oligo dT and Random Hexamer): 500 μl (Yellow Cap)
♦ Nuclease free water: 500 μl (Blue Cap)
This Super RTEasy enzyme is preblended with RNase Inhibitor.
Storage Instruction: -20°C
Reaction set up: The 2x Reaction mix may generate some precipitation after thawing which will not affect the quality of the reaction. In that case, please mix it thoroughly to resuspend and follow the given protocol.
First-Strand cDNA Synthesis Using Super RTEasy Enzyme
♦ Normally, a 20 μl Super RT reaction volume uses 5 to10 ng of total RNA or (1 to 2 ng of mRNA) for the reaction at temperature 42°C. However, depending on the nature of the reaction total RNA or mRNA amounts may be enhanced and a workable temperature can be chosen between 42°C to 55°C.
Add the following components to a nuclease-free microcentrifuge tube
|2x Reaction mix||10 µl|
|Super RT Easy Enzyme Mix||2 µl|
|5 ng to 10ng total RNA OR 1ng to 2ng of mRNA OR more total RNA / mRNA||variable|
|Nuclease free water||variable|
♦ Step 1: Incubate the reaction mix at 25°C for 5 min
♦ Step 2: Incubate at 42°C – 55°C for 30 min
♦ Step 3: Inactivate the reaction by heating at 95°C for 1 min.