FIG. 4.

Low-resolution S1 nuclease mapping of adsA in S. griseus strains (A) and determination of the transcriptional start point of adsA by high-resolution S1 mapping (B). (A) RNA was prepared from cells grown at 28°C for the indicated number of days on solid medium from the wild-type S. griseus IFO13350 (WT), an A-factor-deficient mutant strain (HH1), and an adpA disruptant (ΔadpA). The wild-type strain grew as substrate mycelium (SM) on day 1, as a mixture of aerial and substrate mycelium (AM) on day 2, and as a mixture of aerial hyphae and spores (SP) on day 3. Strains HH1 and ΔadpA grew only as substrate mycelium. (B) RNA prepared from the wild-type cells grown for 3 days on solid medium was used. The arrowhead indicates the position of the S1-protected fragment. The 5′ terminus of the mRNA was assigned to the indicated position because the fragments generated by the chemical sequencing reactions migrate 1.5 nucleotides further than the corresponding fragments generated by S1 nuclease digestion of the DNA-RNA hybrids (half a residue from the presence of the 3′-terminal phosphate group and one residue from the elimination of the 3′-terminal nucleotide) (53).