Anti-DCP2 | mRNA-decapping enzyme subunit 2

Arachis hypogaea, Brachypodium distachyon, Brassica napus, Cannabis sativa, Capsicum annuum, Glycine max, Gossypium sp., Hordeum vulgare, Malus domestica, Manihot esculenta, Medicago truncatula, Nicotiana tabacum, Oryza sativa, Pisum sativum, Populus sp. , Ricinus communis, Solanum lycopersicum, Solanum tuberosum, Sorghum bicolor, Spinacia oleracea, Theobroma cacao, Triticum sp., Vitis vinifera, Zea mays

Approximately 80 mg of Col-0 flowers were subjected to cytosolic and microsomal fractionation, purified according to: Microsomal-type membrane purification Fast and reliable preparation of microsomal-like membranes from plants tissues (Adapted from Abas & Luschnig, Analytical Biochemistry 2010). The final volume of each fraction was adjusted to 100 µL, and 15 µl of each sample were loaded from Arabidopsis thaliana, Col0 (WT), inflorescences. Samples were separated in the cold on 4–15% gradient SDS–PAGE gel (Biorad) and blotted for 90 min at 80 volts, to Immobilon-P, PVDF membrane, 0.45 µm using wet transfer in the cold. Blot was blocked with 30 min at RT in non-fat milk and incubated with anti-DCP2, 1:2000, in PBS 1x, Tween 0.1%, BSA 1%, NaN3 0,2 mM, ON, 4°C, with agitation. The antibody solution was decanted, and the blot was washed 5x 5 min with PBS 1X, Tween 0.1%, Incubation 30 min in PBS 1X, Tween 0.1%, Milk 5%, followed by incubation with a matching HRP-conjugated secondary antibodies at 1h at RT (dilution in PBS 1X, Tween 0.1%, Milk 5%). The blot was washed as above and developed with a following chemiluminescent detection reagent. Note: increasing blocking to 1h/RT and primary antibody incubation 1h/RT, background signal is going to be decreased.

Courtesy Dr. Gregory SCHOTT, ETH Zurich Honggerberg - IMPB Switzerland

Alternative names: AtDCP2; Protein DECAPPING 2, M(7)GpppN-mRNA hydrolase DCP2, Protein TRIDENT