EIN2 | Ethylene insensitive 2
AS12 1865 | clonality: polyclonal | host: rabbit | reactivity: Arabidopsis thaliana
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|Recommended dilution||1 : 2000 (WB)|
|Expected | apparent MW||
|Confirmed reactivity||Arabidopsis thaliana|
|Predicted reactivity||Brachypodium distachyon, Glycine max, Hordeum vulgare, Nicotiana tabacum, Solanum lycopersicum, Oryza sativa, Phtheirospermum japonicum, Populus trichocarpa, Ricinus communis, Sorghum bicolor, Triticum aestivum, , Zea mays, Vitis vinifera|
|Not reactive in||No confirmed exceptions from predicted reactivity are currently known.|
|Selected references||to be added when available, antibody released in April 2015|
40-60 µg of membrane protein (measured with Millipore Direct Detect spec.) isolated following membrane extraction buffer protocol from Dong et al. 2008 (Plant Journal 53(2): 275-286) from 4 day old dark grown Arabidopsis thaliana seedlings, grown on MS plated with ACC or AVG in the plate, extracted with (10 mM Tris-HCl pH 7.5, 150 mM NaCl, 1 mM EDTA, 10 % glycerol, 1 % Triton X-100, and protease ihibitors cocktail from Sigma) were separated 4-20% Bio-Rad precast SDS-PAGE gel and blotted 1h to PVDF membrane using semi-dry transfer. Blots were blocked with 5% milk for 2h at room temperature (RT) with agitation. Blot was incubated in the primary antibody at a dilution of 1: 2 000 overnight at 4°C with agitation. The antibody solution was decanted and the blot was then washed 5 times for 10 min in TBS-T at RT with agitation. Blot was incubated in secondary antibody (anti-rabbit IgG horse radish peroxidase conjugated, from Agrisera AS09 602) diluted to 1:20 000 in for 1h at RT with agitation. The blot was washed as above and developed for 5 min with Bio-Rad ECL according to the manufacturer's instructions. Exposure time was 25 min (to see full length EIN2).
Antibody is suitable for detection of full length EIN2. Bands below 140 kDa are non-specific.
Courtesy of Jennifer Marie Shemansky, University of Maryland, USA
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