SMT1 | Sterol methyltransferase 1
AS07 266 | Clonality: Polyclonal | Host: Rabbit | Reactivity: Arabidopsis thaliana | Marker antibody of integral ER membrane
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|Recommended dilution||1 : 50-1 : 100 (IL), 1 : 500-1 : 1000 (WB)|
|Expected | apparent MW||
|Confirmed reactivity||Arabidopsis thaliana|
|Predicted reactivity||Amborella trichopoda, Brassica napus, Brassica rapa, Capsella rubella, Citrus clementina, Eutrema salsugineum, Glycine max, Glycine soja, Gossypium mexicanum, Medicago truncatula, Populus trichopocarpa, Prunus persica, Ricinus communis, Theobroma cacao, Vitis vinifera|
|Not reactive in||
Hordeum vulgare, Triticum aestivum, Withania somnifera
SMT1 antibody characterization in Western Blot and immunofluorescence labeling: Boutté Y et al. (2010). Endocytosis restricts Arabidopsis KNOLLE syntaxin to the cell division plane during late cytokinesis. EMBO J. 29, 5465-5458.
|Selected references||LaMontagne et al. (2016). Isolation of Microsomal Membrane Proteins from Arabidopsis thaliana. Curr. Protoc. Plant Biol. 1:217-234. doi: 10.1002/cppb.20020.
Yang et al. (2016). Arabidopsis PROTEASOME REGULATOR1 is required for auxin-mediated suppression of proteasome activity and regulates auxin signalling. Nat Commun. 2016 Apr 25;7:11388. doi: 10.1038/ncomms11388.
Yoshimoto et al. (2014). Quality control of plant peroxisomes in organ specific manner via autophagy. J cell Science, August 1, 2014, 127 (15).
Total protein from Col-0 (wild-type) Arabidopsis thaliana were extracted with 50mM HEPES-KOH buffer containing 250 mM sucrose, 5% glycerol, 50 mM NaPP, 1 mM NaMo, 25 mM NaF, 10mM EDTA, 0.5% PVP, 3mM DTT, 1mM PMSF, 10uM Leupeptin & 10nM Calyculin, and then fractionated by ultracentrifugation at 100,000 x gravity for 30 min at 4°C into soluble (S100) and microsomal (P100) proteins as described in LaMontagne et al. (2016). Isolation of Microsomal Membrane Proteins from Arabidopsis thaliana. Current Protocols in Plant Biology 1:1-18. doi: 10.1002/cppb.20020. 30 µg proteins of total, S100 and P100 fractions were denatured at 37°C for 5 min, separated on a 7.5 % SDS-PAGE and blotted 1h to nitrocellulose using tank transfer. Blots were blocked with 1x PBS (from Fisher Scientific BP665-1) + 0.1 %Tween 20 (PBS-T) + 5% milk for 1h at room temperature (RT) with agitation. Blot was incubated in the primary antibody at a dilution of 1: 1000 overnight at 4°C with agitation in 1x PBS-T + 5% milk. The antibody solution was decanted and the blot was rinsed briefly once, then washed four times for 7 min in 1x PBS-T at RT with agitation. Blot was incubated in secondary antibody (anti-rabbit IgG horse radish peroxidase conjugated) diluted to 1:10 000 in 1x PBS-T + 5% milk for 2 hrs at RT with agitation. The blot was washed as above and developed for 4 min with Amersham ECL (RPN2106). Exposure time was 30 seconds and 2 min.
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