Sec21p | Gamma subunit, COP vesicles

GST fusion of a part of recombinant Sec21 of Arabidopsis thaliana Q0WW26, At4g34450

98 kDa

Brachypodium distachyon, Brassica napus, Brassica rapa subsp. pekinensis, Capsella rubella, Citrus clementina, Coffea canephora, Eutrema salsugineum, Glycine max, Glycine soja, Hordeum vulgare var. distichum, Medicago truncatula, Oryza sativa, Populus trichocarpa, Prunus persica, Ricinus communis, Solanum lycopersicum, Solanum tuberosum, Sorghum bicolor, Theobroma cacao, Triticum aestivum, Vitis vinifera, Zea mays,

Application example

50 µg of total protein from (1) Nicotiana tabacum protoplast total protein, (2) Arabidopsis thaliana protoplast soluble protein, (3) Arabidopsis thaliana protoplast total protein were separated on 10 % SDS-PAGE and blotted 2h to nitrocellulose (Semi-dry, 200mA). Filters were blocked over night with 5% low-fat milk powder in TBS and probed with anti-Sec21p antibodies (AS08 327, 1:1000, 1h) and secondary anti-rabbit (1:20000, 1 h) antibody (HRP) in TBS-Tween. Signal was detected with chemiluminescent detection reagent, exposure time was 1 minute.

Protoplasts were extracted in 50mM Tris, 10 mM EDTA and Triton X100, 0.02%.

Immunofluorescence

Immunofluorescence labelling of rabbit anti-SEC21 (gamma subunit of COP vesicles; red) in 5-day-old root epidermal cells of Arabidopsis thaliana expressing ER-mGFP5-HDEL (ER marker; green). The antibody was diluted 1:1000 and the secondary antibody, donkey anti-rabbit CY5-coupled (Jackson ImmunoResearch) was diluted 1:300. The nuclei were stained with DAPI (blue).

Courtesy of Dr. Anna Gustavsson and Dr. Markus Grebe, Umeå Plant Science Centre, Sweden

This antibody can be used as a Golgi marker in immunolocalization and as a marker of COP1 in Western blot.

References describing immunolocalization (IF) and (IG) studies:

Pimpl et al (2000). In Situ Localization and in Vitro Induction of Plant COPI-Coated Vesicles. Plant Cell. 2000 Nov;12(11):2219-36.
Ritzenthaler et al. (2002). Reevaluation of the Effects of Brefeldin A on Plant Cells Using Tobacco Bright Yellow 2 Cells Expressing Golgi-Targeted Green Fluorescent Protein and COPI Antisera. Plant Cell. 2002 Jan;14(1):237-61.

AS10 683 | Anti-HDEL | endoplasmic reticulum retention signa, (100 µg), monoclonal antibodies

AS10 683-25 | Anti-HDEL | endoplasmic reticulum retention signal, (25 µg), monoclonal antibodies

AS08 325 | Anti-Arf1 | ADP-ribosylation factor 1, Golgi marker, rabbit antibodies

Plant protein extraction buffer

Secondary antibodies

Sec21p is a constituent of the COPI vesicle coatomer.