1. DNase treatment of total RNAs
On ice, mix the following:
* RNAs 10ug (not more than 29.5 ul)
* 10 * RTase buffer 5ul
* RQ1 DNase 2ul
* H2O up to 36.5ul
Mix well, spin down and incubate at 37C for 15min.
Inactive the DNase at 65C for 10min, then transfer the tubes on ice.
2. RTase (Synthesis of the first strand cDNAs)
On ice, add the following to the DNase-treated RNAs:
* 0.1M DTT 5ul
* 10mM dNTPs 5ul
* 0.5ug/ul oligo-dT 2ul
* MMLV-RTase 1ul
* RNase-inhibitor 0.5ul
Incubate the tubes 1 hour at 37C then 2min at 92C.
For long storage, store the cDNAs at -80C, otherwise at -20C.
Perform the PCR by using 1 ul of cDNAs for a 50 ul PCR reaction.