14-3-3 GRF | General regulatory element
AS12 2119 | Clonality: Polyclonal | Host: Rabbit | Reactivity: A. platanoides, A. thaliana, C.australis R.Br., L. longiflorum. M. crystallinum, N. tabacum, O. sativa, P. kurroa, P. vulgaris, algae N. oloabundans, Z.mays
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20-28 kDa (depending upon an isoform)
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This antibody does not bind to 14-3-3-like protein D of Nicotiana tabacum.
Left panel: 150 ng of recombinant Arabidopsis thaliana, Physcomitrella patens and Chlamydomonas reinhardtii GRFs (see more details below) were separated on 12 % SDS-PAGE and blotted 1h to PVDF by semi dry blotting (0.8 mA/cm2 for 60 min). Blots were blocked with 2% skimmed milk powder dissolved in TBST (0.1 % Tween 20) for 1h at room temperature (RT) with agitation. Blot was incubated in the primary antibody at a dilution of 1: 1 000 in blocking solution and incubated 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 1:20 000 in for 1h at RT with agitation. The blot was washed as above and developed for 5 min with ECL Prime western blotting detection reagent according to the manufacturer's instructions. Exposure time was 10 minutes.
Right panel: 50 µg of a total cell extract of Arabidopsis thaliana wilde type. Conditions as above, exposure time 30 seconds.
GRF protein designations: At 14-3-3 lambda (GRF6), At 14-3-3 chi (GRF1), At14-3-3 psi (GRF3), At14-3-3 omega (GRF2), At14-3-3 upsilon (GRF5), At14-3-3 omicron (GRF11), At14-3-3 kappa (GRF8), Pp 14-3-3 Pp1s 73_133V6 (closest homolog to At GRF6), Cr 14-3-3 Cre 12.g559250 (closest homolog to AtGRF6). Note that all recombinant GRFs were detected by this antibody, At14-3-3 kappa (GRF8) to a lesser extent.
Courtesy of Dr. Bernhard Wurzinger, University of Vienna, Austria
This antibody is recognizing recombiant GRF of Lilium longiflorum Lil1433_0 accession: AF191746, Lil1433_2, accession: EF397608 and recombinant GRF1,2,3,5 and 6 of Arabidopsis thaliana: GRF1 14-3-3 chi (At4g09000.1), GRF2 14-3-3 omega (At1g78300.1), GRF3 14-3-3 psi (At5g38480.1), GRF5 14-3-3 upsilon (At5g16050.1), GRF6 14-3-3 lambda (At5g10450.2), GRF8 14-3-3 kappa (At5g65430.1), GRF11 14-3-3 omicron (At1g34760.1).
There is also very weak reaction to Physcomitrella patens Pp14-3-3 Pp1s 73_133V6 (closest homolog to AtGRF6) and Chlamydomonas reinhardtii Cr 14-3-3 Cre 12.g559250 (closest homolog to AtGRF6).
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