cDNA sythesis using Vilo from Invitrogen

Overview

This protocol describes how to make cDNA from RNA using Invitrogen's ViLo kit. This kit will make single stranded cDNA using first-strand synthesis from single stranded RNA, normally purified using Trizol. The Vilo kit uses random primers (not oligo DT).

Protocol

• Add ingredients to PCR tube.
• DO NOT FORGET TO MAKE A NO RT CONTROL. Omit the SuperScript Enzyme mix and replace with H2O
• Gently mix ingredients by pipetting up and down.
• Place PCR tubes in a thermal cycler and run the following program
  • 25ºC 10 min
  • 42ºC 60 min to 120 min (Longer reaction time will give you more cDNA)
  • 85ºC 5 min (to terminate reaction)
• If you started with more than 2 µg of cDNA, the cDNA can be diluted 1:20, which will result in a concentration of ~100 ng/µl.
• Store at -20ºC

Reaction mix