ArticleNature protocols2012
Determining proteome-wide expression levels using reverse protein arrays in fission yeast.
Article in Nature protocols, 2012. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 7 papers.
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Who cites it
7 citing papers in PubMed.
- Anticodon Wobble Uridine Modification by Elongator at the Crossroad of Cell Signaling, Differentiation, and Diseases.Epigenomes · 2020Review
- Reciprocal regulation of TORC signaling and tRNA modifications by Elongator enforces nutrient-dependent cell fate.Science advances · 2019Article
- Article
- Article
- Fission yeast Cdk7 controls gene expression through both its CAK and C-terminal domain kinase activities.Molecular and cellular biology · 2015Article
- Synchronized fission yeast meiosis using an ATP analog-sensitive Pat1 protein kinase.Nature protocols · 2014Article
- A coordinated codon-dependent regulation of translation by Elongator.Cell cycle (Georgetown, Tex.) · 2012Article
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Authors and funding
4 authors.
Funding
No grant is acknowledged in the PubMed record.
Abstract
Global protein expression profiling of various mutants or growth conditions is currently a major challenge in biology. Here we provide a protocol for a strategy that we recently developed that couples ORFeome-based (ORF denotes open reading frame) expression to reverse protein arrays; this approach accurately quantifies more than 99% of the predicted fission yeast proteins in various genetic backgrounds. The first stage of this two-stage protocol requires mass mating between any fertile fission yeast mutant of interest and the integrated fission yeast-tagged ORFeome followed by selection of recombinant haploids. The second stage of the protocol, called reverse protein arrays, involves simple large-scale extraction of total proteins, which are then spotted on nitrocellulose membranes for detection by quantitative dot blot. When handled manually, the entire protocol takes about 2 months. However, the process could easily be automated and should also be applicable to other organisms.
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