EMSA of 26 bp P-ABRE (Em1a) with the nuclear extract (NE) prepared from lamina of different rice plants & demonstration of the specificity of binding of DNA-protein complex formation

Copyright information:Taken from "An ABRE-binding factor, OSBZ8, is highly expressed in salt tolerant cultivars than in salt sensitive cultivars of indica rice"BMC Plant Biology 2006;6():18-18.Published online 30 Aug 2006PMCID:PMC1574319. A. 100 fold molar excess of non-radioactive 1X ABRE-DNA (lane 2) or 4X ABRE-DNA (lane 3) or 2X-ABRC-DNA (lane 4) or 4X DRE-DNA (lane5) or 2X ERE-DNA (lane 6) was added in the nuclear extracts from Pokkali plants before the addition of probe and incubated for 30 minutes at 25°C. In lane 1, no competitor was added as control. Equal amount of NE was incubated with amount of probe (1μl = 90,000 cpm) at R.T. in all cases. B. Comparative EMSA of the laminar nuclear extract of two salt sensitive, M-I-48 (lane 2, 3) and IR 72 (lane 4, 5) and two salt tolerant, Pokkali (lane 6, 7) and Nonabokra (lane 8, 9) rice plants with [P]-ABRE-DNA; and the effect of salinity stress to the plants. C. EMSA with nuclear extracts prepared from laminar tissue of control and salt-treated IR-29 (salt sensitive) and IR-8 (moderately salt tolerant) rice cultivars. Histogram drawn from the values obtained by the densitometric scanning of each lane is shown under the autoradiogram. Mean values ± SD of four independent experiments are shown. D. Protein profile of the nuclear extracts (NE) separated by SDS-PAGE followed by staining with Coomassie Brilliant Blue G-250. Molecular weight marker (broad range from New England Biolabs) was loaded in the extreme right side lane.