PPR | Pentatricopeptide repeat-containing protein (chloroplastic) (SOT1)
AS15 3102 | Clonality: Polyclonal | Host: Rabbit | Reactivity: Arabidopsis thaliana
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Total protein (first 2 lanes to the left) or chloroplast protein (2 lanes to the right) of Arabidopsis thaliana were extracted with 100 mM Tris–HCl, pH 8.0, 150 mM NaCl, 1 mM EDTA, 0.5% Triton X-100, 1 mM DTT, 1 mM PMSF, protease inhibitor cocktail, denatured with Laemmli buffer (62.5 mM Tris–HCl pH 6.8, 2% SDS, 10% glycerol, 100 mM DTT, 0.1% β-mercaptoethanol) at 95°C 5 min and separated on 8% SDS-PAGE and blotted 1h to PVDF, using wet transfer. Blot was blocked with 5% milk for 1h/RT or 4°C/ON with agitation. Blot was incubated in the primary antibody at a dilution of 1: 2 000 for 1h/RT with agitation in TBS-T. The antibody solution was decanted and the blot was washed 3 times for 10 min in TBS-T at RT with agitation. Blot was incubated in Calbiochem matching secondary antibody (anti-rabbit IgG horse radish peroxidase conjugated) diluted to 1:25 000 in for 1h/RT with agitation. The blot was washed as above and developed for 10 min with Clarity Western ECL Substrate (Biorad) and visualised with chemiluminescence CCD camera Fluorchem SP (Gel Biosciences). Exposure time was 1-5 minutes.
This antibody is required to be used on chloroplast fraction. There is no signal in the total cell extract.
Courtesty of Dr. Aleksandra Kwaśnik, University of Warsaw, Poland
PPR (Pentatricopeptide repeat-containing protein, chloroplastic - SOT1) is located in chloroplast. Alternative name: Pentatricopeptide repeat-containing protein At5g46580, chloroplastic.
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