RT PCR
Stratagene random oligomers - 8ul
Total RNA ( 1ug/ul) - 4 ul
Put at 70 o for 10 minutes
Ice , and quick spin.
Add: 4ul 1st strand buffer ( BRL)
2ul 0.1 M DTT
1 ul 10 mM each dNTP
Put at 42 o for 2 minutes.
Add 1 ul Superscript II ( 200 units/ ul BRL )
Put at 42 o for 50 minutes.
Quick spin and add 1 ul of RNAse H (2 units / ul BRL )
Put at 37o for 20 minutes.
Use 0.5 to 1 ul to do PCR reaction.