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Alkaline-SDS cell lysis with acetone protein precipitation for proteomic sample preparation of microbes in 96-well plate format

ABSTRACT: Automated proteomic sample preparation offers a solution to the large sample throughput demands in the biotechnology field where hundreds or thousands of engineered microbes are constructed for testing is routine. Meanwhile, sample preparation methods that work efficiently on broader microbial groups are desirable for new applications of proteomics in other fields, such as microbial communities. Here, we detail a step-by-step protocol that consists of cell lysis in alkaline chemical buffer (NaOH/SDS) followed by protein precipitation with high-ionic strength acetone in 96-well format. The protocol works for a broad range of microbes (e.g Gram-negative bacteria, Gram-positive bacteria, non-filamentous fungi) and the resulting proteins are ready for tryptic digestion for bottom-up quantitative proteomic analysis without the need for desalting column cleanup.

INSTRUMENT(S): Orbitrap Exploris 480

ORGANISM(S): Escherichia Coli Rhodotorula Toruloides Np11 Saccharomyces Cerevisiae (baker's Yeast) Pseudomonas Putida Kt2440

SUBMITTER: Christopher Petzold

LAB HEAD: Chris Petzold

PROVIDER: PXD039268 | Pride | 2023-06-23

REPOSITORIES: Pride

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