2d dige labelling buffer  (GE Healthcare)

Journal: Journal of Proteome Research

Article Title: Blue Light-Induced Proteomic Changes in Etiolated Arabidopsis Seedlings

doi: 10.1021/pr500010z

Figure Lengend Snippet: 2D DIGE analysis of the blue light response in the microsomal fraction of 4 day old etiolated Arabidopsis seedlings. (A, B) Col-0 seedlings were irradiated with blue light for 20 min, and microsomal proteins from both control (unirradiated, proteins labeled with Cy3) and irradiated seedlings (proteins labeled with Cy5) were analyzed by 2D DIGE. (A) Superimposed 2D DIGE image of the upper-half of a gel. Proteins (in different modification isoforms) induced by blue light treatment appear as red spots, and those decreased by the treatment appear green, whereas those remaining constant appear yellow. Spots that were characterized by mass spectrometry are highlighted by arrows, and their identities are listed in Table . (B) Zoomed-in 2D DIGE overlay image (from a different gel than that in panel A) showing the blue light-stimulated accumulation of WEB1 in the microsomal fraction. (C) Identities of the row of phot1 spots were further confirmed by 2D DIGE analysis of the microsomal protein of the etiolated seedlings of the phot1–5 (labeled with Cy3, green) and gl (its genetic background, labeled with Cy5, red) mutants. The estimated pH ranges following IEF are indicated above the gel images in Figures – .

Article Snippet: The protein extract was dissolved in 2D DIGE buffer (6 M urea, 2 M thiourea, and 4% CHAPS) and quantified using the Bio-Rad protein assay.

Techniques: Irradiation, Control, Labeling, Modification, Mass Spectrometry

Journal: Journal of Proteome Research

Article Title: Blue Light-Induced Proteomic Changes in Etiolated Arabidopsis Seedlings

doi: 10.1021/pr500010z

Figure Lengend Snippet: Time-dependent phot1 phosphorylation (A) and dephosphorylation (B) after irradiation of dark-grown seedlings. (A) Etiolated Col-0 seedlings were irradiated with blue light for the indicated times, and the microsomal proteins from irradiated (red) and unirradiated samples (green) were compared by 2D DIGE. Shown are the overlay images containing the phot1 region. (B) After saturating irradiation for 20 min, the etiolated seedlings were kept in the dark. The subsequent phot1 dephosphorylation was monitored by 2D DIGE, comparing the microsomal protein from the Col-0 seedling collected at the time points indicated with that of the control samples (collected immediately after irradiation).

Article Snippet: The protein extract was dissolved in 2D DIGE buffer (6 M urea, 2 M thiourea, and 4% CHAPS) and quantified using the Bio-Rad protein assay.

Techniques: Phospho-proteomics, De-Phosphorylation Assay, Irradiation, Control

Journal: Journal of Proteome Research

Article Title: Blue Light-Induced Proteomic Changes in Etiolated Arabidopsis Seedlings

doi: 10.1021/pr500010z

Figure Lengend Snippet: Phosphorylation is the major form of phot1 post-translational modification, as indicated by λ-phosphatase treatment. Phosphatase treatment induced shifts of phot1 spots in microsomal protein from both blue light-irradiated (A) and control (B) seedlings. On the left, “blue and blue/λPP” indicates the comparison of phosphatase-treated (cy5, red) verse untreated (cy3, green) microsomal proteins from blue light-irradiated seedlings by 2D DIGE and “dark and dark/λPP” indicates the comparison of phosphatase treated (cy5, red) verse untreated (cy3, green) proteins from unirradiated seedlings. Proteins that show up in the phosphatase-treated samples appear red, whereas those in the untreated samples are green. Red arrows point to phot1 spots from samples of irradiated seedlings, green arrows point to those from the unirradiated samples, and white arrows point to those from phosphatase-treated proteins.

Article Snippet: The protein extract was dissolved in 2D DIGE buffer (6 M urea, 2 M thiourea, and 4% CHAPS) and quantified using the Bio-Rad protein assay.

Techniques: Phospho-proteomics, Modification, Irradiation, Control, Comparison

Journal: Journal of Proteome Research

Article Title: Blue Light-Induced Proteomic Changes in Etiolated Arabidopsis Seedlings

doi: 10.1021/pr500010z

Figure Lengend Snippet: 2D DIGE Identified phot1 and WEB1 as Blue Light-Responsive Proteins in Arabidopsis a

Article Snippet: The protein extract was dissolved in 2D DIGE buffer (6 M urea, 2 M thiourea, and 4% CHAPS) and quantified using the Bio-Rad protein assay.

Techniques: Sequencing