GPA1 | Guanine nucleotide-binding protein subunit alpha 1 (affinity purified)
AS12 2370 | clonality: polyclonal | host: rabbit | reactivity: Arabidopsis thaliana
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|Recommended dilution||1 : 4000 (WB)|
|Expected | apparent MW||
|Confirmed reactivity||Arabidopsis thaliana|
|Predicted reactivity||Brassica napus, Capsicum annuum , Cynara cardunculus var. scolymus, Eschscholzia californica, Glycine max, Gossypium hirsutum, Hordeum vulgare, Lupinus luteus, Medicago truncatula, Morus notabilis, Nicotiana benthamiana, Nicotiana tabacum, Oryza sativa, Pisum sativum, Phaseolus vulgaris, Populus trichocarpa, Ricinus communis , Solanum lycopersicum, Solanum tuberosum, Sorghum bicolor, Spinacia oleracea, Theobroma cacao, Triticum aestivum, Zea mays, Zostera marina, Vitis vinifera|
|Not reactive in||No confirmed exceptions from predicted reactivity are currently known.|
The nature of a 37 kDa cross-reacting band is not known.
To be added when available, antibody released in April 2016.
25 µg of total protein from the indicated Arabidopsis thaliana wilde type and gpa1-3 mutant were extracted with extraction buffer ( 250 mM sucrose, 100 mM HEPES-KOH pH 7.5, 5% glycerol, 1mM Na2MoO4 x 2H2O, 25mM NaF, 10 mM EDTA, 1mM DTT, 0.5% Triton X-100, protease inhibitor cocktail) and denatured with SDS loading dye (50 mM Tris-HCl pH6.8, 100mM DTT, 2%SDS, 10% glycerol, 0.025% bromophenol blue) at 70°C for 2-5 min were separated on 10% SDS-PAGE and blotted 1h to PVDF using tank transfer. Blots were blocked with Tris-buffered saline containing 0.05% Tween-20 (TBS-T) and 5% skimmed milk powder for 1h at room temperature (RT) with agitation. Blot was incubated in the primary antibody (anti-GPA1, AS12 2370) at a dilution of 1: 5 000 overnight at 4°C with agitation. The antibody solution was decanted and the blot was rinsed briefly twice, then washed 5 times for 15 min in TBS-T with milk powder at RT with agitation. Blot was incubated in secondary antibody (Goat-Anti-Rabbit HRP conjugate, AS09 602) diluted 1:5000 for 2h at RT with agitation. The blot was washed as above with TBS-T without milk powder and then developed with Thermo Scientific SuperSignal West Substrate (pico and femto mixed 1:1), before exposure to a CEA RP NEW film for 10min.
Courtesy of Dr. Elena Petusching, Georg-August-University Goettingen, Germany
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