Fig. 3.

rMRVs with long SIVmac239-gag inserts. (A) Schematics of the in-frame-modified L1 plus-strand RNAs used for creating these rMRVs. See Figs. 1 and 2 for labeling and color schemes. (B and C) Electropherograms of genomic dsRNAs from rMRVs WT (purified virions) and two separately plaque-derived isolates of L1/gag900w/2A/5′66w+M1/3′153 (B) and L1/ gag918w/2A/5′207w/3′168+M1/3′153 (C). Note that for recovering these rMRVs, both modified L1 and modified M1 constructs were used, and the two dsRNA bands corresponding to these slower-migrating, lengthened segments are indicated in both B and C (arrowheads). For C, isolate 1 from B was rerun for comparison; the L1 segment in rMRV L1/gag918w/2A/5′207w/3′168+M1/3′153 is 327 bp longer than the one in L1/gag900w/2A/5′66w+M1/3′153, which explains its even slower migration. (D) Fluorogram of [³⁵S]Met-labeled, in vitro translation products of plus-strand RNAs transcribed from the indicated plasmid constructs of MRV segments. Construct pL1/gag900w/2A/5′66w RNA yields two bands consistent with the expected Gag-containing polypeptide and a putative breakdown product (arrowheads).