FIG. 4.

Primed RNA synthesis on the 8-nt oligoribonucleotide by wild-type primase and mutant proteins. (A) Sequence of the 8-nt oligoribonucleotide primer and G4oric DNA template. The 8-nt oligoribonucleotide 5′-AGUAGGGA-3′ corresponded to the first 8 nucleotides (+1 to +8) of the pRNA product made by primase on the R199/G4oric template. (B) Primed synthesis on γ-³²P-labeled 8-nt oligoribonucleotide (5′-AGUAGGGA-3′) by wild-type and mutant primase proteins in the presence of Mg²⁺ ions. CTP and GTP were added as a source of nucleotides for the elongation reaction. γ-³²P-labeled 11-nt RNA was used as a size marker. The control reaction contained no primase protein (lane 3). (C) Primed synthesis on γ-³²P-labeled 8-nt oligoribonucleotide (5′-AGUAGGGA-3′) by wild-type and mutant primase proteins in the presence of Mn²⁺ ions. CTP and GTP were added as a source of nucleotides for the elongation reaction. γ-³²P-labeled 11-nt RNA was used as a size marker. The control reaction mixture contained no primase protein (lane 12).