Figure 2.

Formation of secondary constriction on mitotic chromosomes is UBF-dependent. (A) Schematic representation of the novel secondary constriction formed by the insertion of a 1.4-Mb XEn array into the q-arm of chromosome 10 in pseudo-NOR clone 3D. (B) Western blot of proteins extracted from pseudo-NOR clone 3D cells transfected with control or UBF siRNAs revealed a 13-fold depletion of UBF1 and UBF2. (C) Metaphase chromosome spreads from pseudo-NOR clone 3D were hybridized with an XEn probe. Arrowheads indicate the site of pseudo-NOR insertion. Transfection of UBF siRNA induces the loss of pseudo-NOR secondary constriction. (D) Combined FISH and silver staining of metaphase spreads from pseudo-NOR clone 3D. UBF depletion results in the loss of silver staining at pseudo-NORs (arrowheads). Note the silver staining of an endogenous NOR (*) in the bottom right panel.