Figure 3.

Subcellular Fractionation of GFP-αTIP Fusion Proteins Expressed in Protoplasts.

Protoplasts were transfected with no DNA (lanes 1 and 2), with GFPαTIP expression plasmid DNA (wt, lanes 3 and 4), or with GFPαTIPΔ expression plasmid DNA (Δ, lanes 5 and 6). Seventeen hours later, protoplasts were harvested and separated into pellet (P) and vacuole (V) fractions. Aliquots representing 10% of each fraction were separated on a 4 to 20% acrylamide gradient SDS-PAGE gel, transferred to nitrocellulose, and α-TIP protein detected with a 1:2000 dilution of anti-α-TIP protein antiserum. Arrows, full-length fusion protein; asterisk, ∼23-kD proteolytic product specific for GFPαTIPΔ. Positions of molecular mass markers are indicated in kilodaltons at right.