FIG. 3.

Schematic for generation of viral transcripts containing mouse-adapted amino acid changes in either the 2B or 3A protein. A DNA copy of the HRV39 and RV39/L viral genomes was cloned into pACYC177. The plasmids were cleaved with BsiEI and SacI or BsiEI and SphI, and DNA fragments containing (i) the T7 promoter and the 5′ half of the viral cDNA or (ii) the 3′ half of the viral cDNA and the poly(A) sequence were purified. Ligations of the 5′ half of HRV39 DNA and the 3′ half of RV39/L DNA or the 5′ half of HRV39/L DNA and the 3′ half of HRV39 DNA resulted in full-length DNAs with changes in either 2B or 3A. RNA transcripts were generated by in vitro transcription, and the RNA was introduced into cells by transfection to produce virus.