cDNA Synthesis with ThermoScript
Dominik Haudenschild From Gibco cat11146-024, March 12, 2001


Anneal Random Primers
up to 9 µl total RNA (10pg to 5µg)
1µl Random Primers
Water to 10µl

Heat to 65°C for 5 minutes to open secondary structure
Place on Ice

Prepare Master Mix
4µl 5x cDNA Synthesis Buffer
1µl 0.1M DTT
1µl RNaseOUT
1µl DEPC Water
2µl 10mM dNTP mix
1µl ThermoScript Reverse Transcriptase

cDNA Synthesis
Add 10µl of Master Mix to each tube of RNA
Place in PCR machine programmed to:
25°C for 10 minutes
55°C for 50 minutes
85°C for 5 minutes

Add 1µl RNase-H and incubate at 37°C for 20 minutes (optional)
Store -80 or use immediately for PCR