EF1A | Elongation factor 1-alpha / EF-1-alpha

Full length, recombinant EF1A of Arabidopsis thaliana UniProt: P0DH99-1, TAIR: AT1G07940

application example

10 µg total protein extracted from 15 day old Arabidopsis thaliana seedlings that were either kept at 22°C (3) or heat treated at 38°C for 2 hours  prior to protein extraction (4). As positive control 10 ng of recombinant AtEF1A (2) were separated side by side with the plant samples on 10-15 % gradient SDS-PAGE and blotted to nitrocellulose (Bio-rad). Blots were blocked following transfer with 5% low fat milk in low salt buffer for 1h at room temperature with agitation. Blots were incubated in the primary antibody at a dilution of 1: 1000 for O/N 4°C with agitation in the blocking solution. The primary antibody solution was removed and the blot was rinsed briefly twice, then washed 4 times for 15 min each at room temperature with agitation using low salt buffer. Blots were incubated in secondary antibody, anti-rabbit IgG horse radish peroxides conjugated, from Agrisera, AS09 602, diluted to 1:35 000 for 1h at room temperature with agitation then washed as above and treated with ECL detection reagent according to the manufacturer's instructions.  Exposure time was 5 seconds.

Lane (1) with no signal: recombinant Triticum spp. EF1A.

Courtesy Dr. Eman Basha, Arizona State University

EF1A (elongation factor 1-alpha) belongs to the GTP-binding elongation factor family and is localized in the cytoplasm. It is an essential enzyme in elongation phase of protein synthesis. There are four genes encoding EF1A in Arabidopsis, sequences of genes 1,2 and 3 are identical. Synonymes:eEF-1A.