Part of my research involves amplification of some rather large cDNAs (6-7kb) in several different plant species. If you are familiar with the 5’ RLM RACE kit from Ambion and I was hoping you may be able to help me with a technical question.
I’ve decided to use the RLM RACE kit from Ambion, but since we already have a lot of the components, I’m ordering the primers and reagents required for the protocol. The kit uses a 5’ RACE Adapter RNA primer which is ligated to the de-capped mRNA. I’m wondering if I might be able to avoid ordering a terribly expensive primer (RNA oligo) by just specifying a conversion of the 3’ nucleoside to an RNA to enable the ligation (with the rest of the nucleosides being DNA). I thought the RT enzyme would not reverse transcribe the RNA-DNA hybrid at the cap. Do you think this approach would work?
The 5’ adapter primer sequence is 5’- GCUGAUGGCGAUGAAUGAACACUGCGUUUGCUGGCUUUGAUGAAA-3’
I would very much appreciate any thoughts or suggestions you have on this issue.