Figure 12.

Isolation of PVCs from BY-2 Cells.

(A) Identification of fractions enriched in VSR proteins. Cell homogenates were layered onto a discontinuous sucrose density gradient (20 and 60% [w/w]). Fractions were collected and subjected to SDS-PAGE followed by immunodetection with VSR antibodies. M, molecular marker in kilodaltons.

(B) The VSR-eniched fractions from (A) (fractions 6 to 8) were further layered onto a continuous sucrose density gradient (25 to 50% [w/w]) and centrifuged isopycnically. Fractions were collected for protein gel blotting using various antibodies.

(C) Progressive enrichment of VSRs in subcellular fractionation. Total protein from BY-2 cell homogenate (lane 1), VSR-enriched fractions 6 to 8 of (A) (lane 2), and fraction 11 from (B) (lane 3) were used for protein gel blot analysis using various antibodies. Equal amounts of protein (15 μg) applied to each lane.