Flanked by Gateway recombination cassettes (Supplemental Table S4) and cloned intoMurcha et al.pDONR201. LR reactions were carried out with C-terminal GFP fusion vectors for GFP localization (Carrie et al., 2009), pDEST14 for in vitro transcription and translation, and pB2GW7 (35S CaMV promoter) for Agrobacterium tumefaciensmediated complementation (Karimi et al., 2002). Yeast two-hybrid vectors w
Flanked by Gateway recombination cassettes (Supplemental Table S4) and cloned intoMurcha et al.pDONR201. LR reactions were carried out with C-terminal GFP fusion vectors for GFP localization (Carrie et al., 2009), pDEST14 for in vitro transcription and translation, and pB2GW7 (35S CaMV promoter) for Agrobacterium tumefaciensmediated complementation (Karimi et al., 2002). Yeast two-hybrid vectors w
Mitochondrial pellet was resuspended in digitonin buffer (30 mM HEPES-KOH, pH 7.4, 150 mM potassium acetate, 10 [v/v] glycerol, and 1 mg of digitonin) and kept on ice for 30 min. Ten microliters of antibody serum, 50 mL of Protein A-Sepharose, 1 (w/v) BSA, and Complete protease inhibitor (Roche) were added to the mitochondria and incubated for 6 h at 4 on a rotating wheel at 20 rpm. Beads were
On kit (Promega); 10 mL of radiolabeled protein was added to the reaction, and samples were resolved by SDS-PAGE, dried, and exposed to a phosphor-imaging screen.Plant Material and Growth ConditionsArabidopsis (Arabidopsis thaliana) T-DNA insertional knockout lines for Tric1 (SALK_031707, SALK_112126, and At3g49560) and Tric2 (SALK_136525, SALK_149871, and At5g24650) were obtained from ABRC and sc
On kit (Promega); 10 mL of radiolabeled protein was added to the reaction, and samples were resolved by SDS-PAGE, dried, and exposed to a phosphor-imaging screen.Plant Material and Growth ConditionsArabidopsis (Arabidopsis thaliana) T-DNA insertional knockout lines for Tric1 (SALK_031707, SALK_112126, and At3g49560) and Tric2 (SALK_136525, SALK_149871, and At5g24650) were obtained from ABRC and sc
Mitochondrial pellet was resuspended in digitonin buffer (30 mM HEPES-KOH, pH 7.4, 150 mM potassium acetate, 10 [v/v] glycerol, and 1 mg of digitonin) and kept on ice for 30 min. Ten microliters of antibody serum, 50 mL of Protein A-Sepharose, 1 (w/v) BSA, and Complete protease inhibitor (Roche) were added to the mitochondria and incubated for 6 h at 4 on a rotating wheel at 20 rpm. Beads were
L cycle regulation. The LxCxE motif and the tripeptide HDP of the J domain of the polyomavirus tumor proteins make direct contacts with the Rb proteins and Hsc70, respectively; together they effect the release of members of the E2F family of transcription factors from their Rb partners to promote cell cycle progression [reviewed in 55]. There is less certainty about the identity of tAg amino acids
L cycle regulation. The LxCxE motif and the tripeptide HDP of the J domain of the polyomavirus tumor proteins make direct contacts with the Rb proteins and Hsc70, respectively; together they effect the release of members of the E2F family of transcription factors from their Rb partners to promote cell cycle progression [reviewed in 55]. There is less certainty about the identity of tAg amino acids