Figure 1.

Hybridization of the 225- and 180-kb amplicons with cosmids representing the telomere-proximal third of NotI fragment L (∼120 kb). (A) The subregion of NotI fragment L that hybridizes with the amplicons and characteristics of the amplified DNAs are shown. NotI fragment L is 380 kb long, and approximately one-third of it hybridizes to the amplicons. TEL, telomere; TAS, telomere-associated sequences; LTR, long terminal repeat. The cosmids, which are numbered, are drawn at their approximate locations. The sod2 gene is depicted as a rectangle. The centromere-proximal 5 kb of hybridization between wild-type genomic DNA and the 180-kb amplicon contains sod2, a long terminal repeat sequence, a stretch of 30 adenosine residues, and two ORFs. The centromere-proximal 1 kb of hybridization between wild-type genomic DNA and the 225-kb amplicon contains a 134-bp inverted repeat and a 290-bp spacer. (B) Wild-type chromosomal DNA digested with NotI. The digest was separated by pulsed-field gel electrophoresis. Left, ethidium bromide–stained gel. Right, hybridization with labeled 225-kb amplicon. NotI fragment L contains sod2 and a telomere. NotI fragments C, I, and M all contain telomeres. A indicates intact chromosome III. NotI fragments C and A do not separate well under the running conditions used in this experiment. (C) Cosmids from the Mizukami library (Mizukami et al., 1993) digested with HindIII, with the most telomere-proximal cosmid on the left. Left, ethidium bromide staining. Right, duplicate transfers hybridized with the 225- or 180-kb amplicon. All of NotI fragment L was tested, and only cosmids in the region shown hybridized. DNA Marker X (Boehringer Mannheim) was used for size markers.