AKIN10 | SNF1-related protein kinase catalytic subunit alpha KIN10
AS10 919 Clonality: Polyclonal Host: Rabbit | Reactivity: Arabidopsis thaliana
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Oryza sativa, Solanum lycopersicum, Vitis vinifera
Arabidopsis thaliana total proteins were extracted in Hepes buffer (50mM Hepes-NaOH pH 7.8; 2mM EDTA pH 8.0; 1mM DTT; Phosphatase and Protease inhibitor cocktails). After centrifugation, the supernatant was recovered and protein concentration determined using the Bradford protein assay. 5, 10, 20 and 30 μg of total protein were resolved, for each plant extract, by SDS-PAGE, transferred to a PVDF membrane and analyzed by immunoblotting with α-AKIN10 antibody (Agrisera; AS10 919; 1:500; 1% non-fat Milk in TBS O/N at 4°C). Secondary antibody anti-rabbit HRP-conjugated was used at 1:20 000, 1% non-fat Milk in TBS for 2h at RT. Washes were made as following: after blocking, the membrane was washed 2X 5 min with 1x TTBS; after the primary antibody incubation, the membrane was washed 5x 5 min with 1x TTBS; after the secondary antibody incubation, the membrane was washed 5x 5 min with 1x TTBS (contained Tween-20 at 500 µl/liter), followed by 3x 5 min with 1x TBS Wash Buffers (TBS and TTBS): 1X TBS (1 liter) 2.42 g Tris 8 g NaCl Adjust pH to 7.5-7.6 with HCl 1X TTBS: 1X TBS + 500 ul Tween-20 per liter. Reaction was developed following manufacture's recommendations and recorded using BioRad GelDoc.
Courtesy of Drs. Leonor Margalha/Elena Baena-González, Instituto Gulbenkian De Ciencia, Portugal
AKIN10 (E.C.= 22.214.171.124) is a catalytic subunit of the putative trimeric SNF1-related protein kinase (SnRK) complex, which may play a role in a signal transduction cascade regulating gene expression and carbohydrate metabolism in higher plants. Synonymes: AKIN alpha-2
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