Figure 13.

Exonuclease I (Exo-I) efficiently remove single-stranded oligos after liquid hybridization. Extracted RNA1 was hybridized with either biotin-labeled GAPDH double-stranded (ds) probe (lanes 1–7) or single-stranded biotinylated primer probe (lanes 8–11) using the liquid hybridization method. After completion of hybridization, the samples were divided into half and either treated with Exo-I overnight at 37 °C or left at 4 °C without Exo-I. Both the Exo-I-treated and untreated hybridized samples were electrophoresed on 1.2% non-denaturing agarose gels in 1× MOPS buffer without formaldehyde followed by transfer onto positively-charged nylon membranes using the semi-dry method. (a) Schematic diagram showing hybridization of RNA and double-stranded labeled probe in the liquid hybridization method. Treatment with Exo-I resulted in the same size of the product after hybridization. (b) Agarose gel image after ethidium bromide staining of the hybridized samples taken by the Typhoon imaging system before transfer to a nylon membrane. (c) X-ray detection of biotin-labeled probe/ product after transfer.