Reverse transcription using nuclease resistant primers

Recent developments in nucleotide chemistry have allowed the synthesis of RNA oligonucleotides modified at the 2'-hydroxyl position on the ribose. The oligo[2'-O-allylribonucleotide], (2'-O-allyl oligo), has been shown to be resistant to a wide array of nucleases (1). Such oligos annealed to RNA form hybrids with a Tm higher than the conventional RNA duplex (2). Furthermore, these hybrids are fully stable in the presence of RNase H. Here we describe the utility of 2'-O-allyl oligos as primers for reverse transcription reactions. A schematic drawing of primers, template mRNAs as well as the predicted sizes of the reverse transcription products are shown in Figure 1. Figure 2 shows that a 2'-O-allyl oligo, A, (lane 1) functions extremely poorly as a primer for the reverse transcription, whereas a DNA oligo of identical sequence, D, (lane 2) and the internal control DNA oligo (eALAS) are extende