Journal: The Journal of Experimental Medicine
Article Title: Mutations in STN1 cause Coats plus syndrome and are associated with genomic and telomere defects
Figure Lengend Snippet: Mutations in STN1 result in abnormal telomere phenotypes. (A) DNA samples, prepared from PBLs of patient P1, her heterozygous father (F1), and a noncarrier sibling (S1) and patient P2, his heterozygous mother (M2), and two independent control samples (C), were analyzed by in-gel hybridization. Duplicated lanes were electrophoresed in the same gel, and then separated and hybridized to a G-rich or C-rich telomeric probe, as indicated above the panels. After native hybridization to detect single-stranded telomeric DNA (top), the gels were denatured and rehybridized with the same probes to detect the overall duplex telomeric DNA (bottom). Treatment with exonuclease I is indicated above the lanes. (B) Graphic illustration of the mean telomere length for the patients and their family members, calculated based on the following number of independent measurements of four in-gels and two Southern analyses: P1:6, M1:3, F1:3, S1:3, P2:9, M2:3, F2:1, C1:2, and C2:3. (C) Graphic illustration of the relative native (single strand) per denatured (total) telomeric signal, normalized to the controls. The values represent the mean of four independent measurements for P1 and four for P2. (D, top) A metaphase spread from P2 PBL after CO-FISH. Bar, 5 µm. The area designated by the white frame in enlarged in the bottom panel. Arrowheads point to chromosome ends with hybridization signals on both sister chromatids, indicating that T-SCE occurred at that chromosome end. (right) The frequency of T-SCE in P2 PBLs is compared with PBLs from four age-matched controls (con-BL1-4). For P2 PBLs, 1,150 chromosome ends were analyzed. For control PBLs, between 600 and 930 telomeres were analyzed (*, P
Article Snippet: Exonuclease I treatment was performed overnight at 37°C with 200 U exonuclease I (Thermo Fisher Scientific) per 5 µg DNA in 500 µl, followed by ethanol precipitation.
Techniques: Hybridization, Fluorescence In Situ Hybridization