10.2.8.3. Column purification of cDNA
The cDNA can also be purified to remove unincorporated tagged-cDNA primer, using Qiagen affinity purification columns, and thus significantly reduce the chance of falsely detecting the incorrect strand through participation of residual tagged-cDNA primer in the early PCR reactions. This procedure is a common alternative to Exonuclease-I digestion (Peyrefitte et al., 2003; Carrière et al., 2007) and used in strand-specific detection of several honey bee viruses (Boncristiani et al., 2009; DiPrisco et al., 2011; Peng et al., 2012).
1. Follow Qiagen
DNA affinity column purification protocol.
2. Elute the purified cDNA in 100 µl nuclease-free water.