Fig. 1. Ldb1 biotinylation and streptavidin pulldown. (A) Schematic representation of bio-Ldb1. The 23 amino acid sequence recognised by BirA and a triple haemagglutinin tag are fused in tandem to the amino-terminal end of the ATG-less Ldb-cDNA. (B) Expression of Ldb1 and bio-Ldb1 in nuclear extracts of induced C88BirA/bio-Ldb1, C88-/BirA and C88^-/- cells. Eight of 16 transfectants are shown. Three clones (lanes 3-5) did not express bio-Ldb1, lane 9 not loaded. Clone #3F4 was chosen for further experiments. Lanes on the right are C88-/BirA and C88^-/- controls. (C) Ldb1 and bio-Ldb1 expression in equal amounts (see Coomassie stained gel) of nuclear extracts of noninduced (-) and induced (+) #3F4 cells. Expression of endogenous Ldb1 is reduced in induced cells. (D) PAGE of proteins bound to bio-Ldb1. Pulldowns of noninduced and induced cell extracts washed at lower stringency conditions are shown. PD, pulled down proteins; NE, untreated nuclear extract.