S14 | 40S ribosomal protein S14-1

PRODUCT INFORMATION IN PDF

application example

20 µg of total protein from Arabidopsis thaliana extracts from (1) total cell, nuclei (2), chloroplasts (3), thylakoids (4) were extracted with preparation buffer (330 mM sorbitol, 25mM Tricine pH 7.8, 1mM EDTA, 10mM KCl, 0,15% BSA, 4mM Na ascorbat and 7mM L-cysteine) and separated on 12 % sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and blotted 1h 30min to Immobilon™PVDF (Millipore) membrane. Blots were blocked with 10% dry milk in Tris-buffered saline TBS-T (50 mM tris, 150 mM NaCl, pH 7.6 + 1ML 20% TWEEN 20) for 1h at room temperature (RT) with agitation. Blots were incubated in the primary antibody at a dilution of 1: 2 000 (in TBS-T) overnight at 4ºC with agitation. The antibody solution was decanted and the blot was rinsed briefly twice, and then washed once for 15 min and 3 times for 5 min in TBS-T at RT with agitation. Blot was incubated in secondary antibody (anti-rabbit IgG horse radish peroxidase conjugated, from Pierce) diluted to 1:20 000 in TBS-T for 1h at RT with agitation. The blot was washed as above and developed for 5 min with ThermoSuper Signal WestPico according to the manufacturers instructions. Exposure time was 3 minutes. Courtesy of Dr. Rikard Fristedt, UCLA, USA

5 µg of total protein from Arabidopsis thaliana (1), Zea mays (2), Chlamydomonas reinhardtii (3), Salmo salar (4)  extracted with PEB  were separated on 4-12 % SDS-PAGE and blotted 1h to PVDF. Blots were blocked  for 1h at room temperature (RT) with agitation. Blot was incubated in the primary antibody at a dilution of 1: 1 000 for 1h at RT with agitation. The antibody solution was decanted and the blot was rinsed briefly twice, then washed once for 15 min and 3 times for 5 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 1:25 000 in  for 1h at RT with agitation. The blot was washed as above and developed for 5 min with ECL according to the manufacturers instructions. Exposure time was 2 minutes.

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Plant and algal protein extraction buffer

Secondary antibodies