FIG. 5.

(A) Details of substitution mutations within the internal palindrome and around the nick site of the MVM insert in pR1-4AGA. Letters indicating wild-type nucleotides are black, and those indicating mutations are white. Mutant 1 has a C-to-G switch at the position in the inboard arm of the internal palindrome which would base pair with G5042 in the cruciform structure. In mutant 2, NS1-less, the core of all possible NS1 binding sites is destroyed by switching every second nucleotide between strands of the cruciform. In mutant 3, son-of-NS1-less, only the binding site core in the inner arm of the palindrome is mutated, by switching three pairs of alternating bases along the strand. Mutant 4 has alternating nucleotides through the NS1 binding site switched between strands. Mutant 5 has the entire consensus nick site deleted; the juxtaposed vector sequences are shown in white letters. (B and C) Autoradiographs of total replication products (B) and anti-NS1 immunoprecipitates of replication products (C) generated using substrates 1 through 5, with each lane numbered according to the substrate used. WT, wild type.