Summary
A major challenge in the field of proteomics is obtaining high quality peptides for comprehensive proteome profiling by liquid chromatography mass spectrometry for many organisms. Here we evaluate and modify a range of sample preparation methods using photosynthetically active Arabidopsis leaf tissues from several developmental timepoints. We find that inclusion of FASP-based on filter digestion improves all protein extraction methods tested. Ultimately, we show that a detergent-free urea-FASP approach enables deep and robust quantification of leaf proteomes. For example, from 4-day-old leaf tissue we profiled up to 11,690 proteins from a single sample replicate. This method should be broadly applicable to researchers working on difficult to process samples from a range of plant and non-plant organisms.
- Chloro
- Methanol/Chloroform Extraction
- FASP
- Filter Aided Sample Prep
- GO
- Gene Ontology
- IAA
- Iodoacetamide
- LFQ
- Label Free Quantification
- MS/MS
- Tandem mass spectrometry
- TF
- Transcription Factor
- UA
- Urea Extraction
- 1D
- 1 Dimensional
- 2D
- 2 Dimensional