Figure 3.
Genotypic characterization of clade 1 Ohio/03 and clade 2 Rich/07 LAIVs: (A) Schematic representation of the hemagglutinin (HA; left) and neuraminidase (NA; right) segments from Ohio/03 (black) and Rich/07 (red) LAIVs, indicating the Sac I (Ohio/03) and Aat II (Rich/07); and the Bgl II (Ohio/03) and Hind III (Rich/07) unique restriction sites and their location in the viral HA and NA, respectively. (B,C) Madin-Darby canine kidney (MDCK) cells (6 well-plate format, 1 × 106 cells/well) were individually infected (MOI 0.1) with Ohio/03 or Rich/07 LAIVs and incubated at 33 °C. At 48 h post-infection (p.i.), total RNA was extracted and the HA (B) and NA (C) viral segments were amplified by RT-PCR using specific primers. Undigested and Sac I- or Aat II-digested PCR products of HA (B) and Bgl II- or Hind III-digested PCR products of NA (C) are shown. M: Ladder marker. The nucleotide size of the different bands of the ladder marker is indicated on the left. Nucleotide length of undigested and digested products for Ohio/03 (black) and Rich/07 (red) are indicated on the right.