FIG. 1.

(A) Sequence homology among the 3′ end of the leader RNA, constructed IGS7/8, and naturally occurring IGS6/7 and IGS5/6 of MHV-A59. The bracket marks the nine-nt core consensus IGS motif. For each IGS, the stop codon of the upstream gene is underlined. (Note that the upstream nonhomologous C residue in IGS6/7 is a U residue in MHV-JHM.) (B) Construction of transcription vectors for the synthesis of MHV DI RNAs containing a new IGS (IGS7/8) downstream of the N gene. The parent vector, pB36, encodes a DI RNA consisting entirely of MHV components, except for a 48-nt linker that connects the 5′ 467 nt of the MHV genome (leader plus gene 1 fragment) to a 1,666-nt segment comprising the entire N gene and 3′ UTR (26). Plasmid pBL83 was derived from pB36 by the insertion of IGS7/8 (hatched rectangle), followed by an 18-bp polylinker region (solid rectangle), immediately downstream of (and partially overlapping with) the N gene stop codon. The PCR primers used in the construction of pBL83 are indicated above the schematic of the relevant segment of this plasmid, and the sequence of the insertion is shown beneath it. The MHV core consensus IGS sequence is boxed, and the N gene stop codon is marked with stars.