RPS6A | 40S ribosomal protein S6-1
AS19 4292 | Clonality: Polyclonal | Host: Rabbit | Reactivity: Arabidopsis thaliana
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0.5 µg/well of total protein extracted freshly from Arabidopsis thaliana whole leaf with extraction buffer ( 50mM Tris-HCl pH 7.5, 1mM NaCl, 1% Triton X-100, 1mM DTT, 1mM PMSF, 0.5mM EDTA and 1X Halt protease and phosphatase inhibitor cocktail) and denatured with laemmli sample buffer at 70°C for 5 min. Protein extract were separated on 4-12% SDS-PAGE and blotted 1h to nitrocellulose (pore size of 0.45um), using wet transfer. Blot was blocked with 5% BSA for 1h/RT with agitation. Blot was incubated in the primary antibody at a dilution of 1: 2000 in 5% BSA+PBS-T ON/4°C with agitation. The antibody solution was decanted and the blot rinsed, then washed 3 times for 10 min in PBS-T at RT with agitation. Blot was incubated in Agrisera matching secondary antibody (anti-rabbit IgG horse radish peroxidase conjugated AS09 602) diluted to 1:25 000 in 5% BSA+PBS-T for 1h/RT with agitation. The blot was washed as above and developed for 5 min with Agrisera ECLBright. Exposure time was 10 seconds.
Courtesy Dr. Irabonosi Obomighie, University of Essex, United Knigdom
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