FIG. 9.

Spb1p is eluted with Nop1p on a Mono-S column. Cellular extracts containing HA-Spb1p (YDK14-1A/pDK376) were fractionated by size chromatography on a Sephadex column. A high-molecular-weight fraction containing HA-Spb1p was then loaded onto a cation-exchange column (Mono-S). Elution was then performed with a gradient of NaCl that was applied in two steps. Fractions of 1 ml were collected, and a sample of each fraction was subjected to an SDS–10% PAGE, transferred to nitrocellulose, and probed with antibodies revealing HA-Spb1p (upper panel), Nop1p (middle), or the ribosomal protein Qsr1 (lower panel). Load, the fraction that was applied onto the Mono-S column; FT, flowthrough fraction that was not retained by the column; lanes 1 to 15, first slope of the gradient from 0 to 0.5 M NaCl; lanes 16 to 23, second slope of the gradient from 0.5 to 1 M NaCl.