Sec8 | Exocyst complex component SEC8

application example
 

50 µg of total protein from Arabidopsis thaliana, wt whole plant extract of one week old seedlings (1), whole plant extract of one week old seedlings of Arabidopsis thaliana Col-0 expressing SEC8-GFP (2), whole plant extract of one week old seedlings of Arabidopsis thaliana sec8 k.o. mutant complemented by SEC8-GFP (3), extracted with SEC6/8 buffer (20mM HEPES, pH 6.8; 150mM NaCl; 1mM EDTA; 1mM DTT; 0.5% Tween 20) were separated on 10 % SDS-PAGE then using semi-dry transfer blotted 1h to the nitrocellulose membrane (1.5 mA/cm2). Blots were blocked with 5 % non fat milk in PBS overnight at 4°C with agitation. Blot was incubated in the primary antibody in 5 % milk in PBS at a dilution of 1: 7500 for 1h at RT with agitation. The antibody solution was decanted and the blot was rinsed five times 5 min with PBST (PBS + 0.5% Tween 20) at RT with agitation. Blot was incubated in secondary antibody (anti-rabbit IgG horse radish peroxidase conjugated) diluted to 1:15 000 in 5 % milk in PBS for 1h at RT with agitation. The blot was washed as above and developed for 5 min with ECL according to the manufacturer's instructions. Exposure time was 10 seconds.

Courtesy of Dr. Michal Hála, UEB, Czech Republic

Sec8 is a member of the exocyst complex involved in tethering vesicles to the plasma membrane during regulated or polarized secretion. Involved in secretory process during cell plate initiation during cytokinesis. Participates in polarized pectn delivery.