CDNA synthesis

Protocol 7.

Method

1. Pipette 5 of total RNA into a 0.5 ml microcentrifuge tube and bring the total volume to 24.5 |xl using DEP-treated water.

2. Briefly heat (95°C, 3 min) and then quickly cool on ice.

3. Add the following reagents.

5 X first strand buffer (GIBCO-BRL), 20 |xl RNAsin (Promega) 20 units, 0.5 (jul random hexamers (62.5 A26o units/ml), 4 |xl dNTPs (2.5 mM each), 40 jjU 0.1 M DTT, 10 ijlI

Superscript™ reverse transcriptase (GIBCO-BRL) (200 units), 1 jjlI

5. Heat at 95°C for 5 min to terminate reactions.

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