Diagram of the ribo-SPIA process for the synthesis of sscDNA. First strand cDNA is generated from template RNA using reverse transcriptase (RT) and two types of chimeric primers, random and oligo(dT), containing an RNA overhang (Step 1). DNA Polymerase is added to the reaction to generate second strand cDNA (Step 2). sscDNA is amplified from the dscDNA template in a cycle in which a SPIA™ primer (DNA/RNA hybrid) anneals to the template, DNA Polymerase begins duplicating the cDNA, the RNA portion of the primer degraded by RNase H (which only degrades RNA when it is in a duplex with DNA), thus allowing another SPIA™ primer to bind to the template and restart the reaction (Step 3).
Watson et al. BMC Genomics 2008 9:84 doi:10.1186/1471-2164-9-84