Proteomics

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Arabidopsis thaliana whole-proteome BIN2 kinase assay


ABSTRACT: Finely ground leaf tissue of wild-type columbia-0 ecotype was used. Protein extraction was performed in buffer-saturated phenol and high sucrose buffer. Extracted proteins were incubated with purified recombinant GST or GST-BIN2 kinase. Filter-assisted sample preparation (FASP) columns in presence of 8M urea were used to further purify proteins. Purified proteins were reduced with TCEP, alkylated with iodoacetamide and digested into peptides with trypsin and lys-C. Digested peptides were desalted with c18 columns and labelled with 11-plex Tandem Mass Tag (TMT11) reagents. Phosphoenrichment was performed in a two-steps tandem pipeline: High-Select TiO2 Phosphopeptide Enrichment kit (Thermo Scientific) was used first and High Select Fe-NTA Phosphoptide Enrichment kit (Thermo Scientific) was used on the flow-through obtained from first enrichment.

INSTRUMENT(S): Q Exactive Plus

ORGANISM(S): Arabidopsis Thaliana (ncbitaxon:3702)

SUBMITTER: Justin Walley  

PROVIDER: MSV000086462 | MassIVE | Tue Nov 17 07:41:00 GMT 2020

REPOSITORIES: MassIVE

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Publications

Integration of multi-omics data reveals interplay between brassinosteroid and Target of Rapamycin Complex signaling in Arabidopsis.

Montes Christian C   Wang Ping P   Liao Ching-Yi CY   Nolan Trevor M TM   Song Gaoyuan G   Clark Natalie M NM   Elmore J Mitch JM   Guo Hongqing H   Bassham Diane C DC   Yin Yanhai Y   Walley Justin W JW  

The New phytologist 20220811 3


Brassinosteroids (BRs) and Target of Rapamycin Complex (TORC) are two major actors coordinating plant growth and stress responses. Brassinosteroids function through a signaling pathway to extensively regulate gene expression and TORC is known to regulate translation and autophagy. Recent studies have revealed connections between these two pathways, but a system-wide view of their interplay is still missing. We quantified the level of 23 975 transcripts, 11 183 proteins, and 27 887 phosphorylatio  ...[more]

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