Data of drought resistance mechanism of Lonicera Hypoglauca
Description
This dataset encompasses all supplementary materials, including physiological and proteomic data obtained during drought and rehydration treatments. The physiological data are presented in Table A.1, while the proteomic data are summarized in Tables A.2-A.5, organized according to the following categories: all 949 differentially expressed protein annotation files, the 96 proteins with the top 7 of KEGG enrichment coefficients, PRM validation results, and 40 differentially expressed protein associated with calcium and phosphorylation regulation. The subcellular localization and KOG classification of the 949 proteins, as well as the KEGG enrichment analysis results for each treatment condition, are provided in Figures A.1 to A.3.
Files
Steps to reproduce
The branches of 5-year-old L. hypoglauca were used for layering propagation. After rooting, the complete seedlings produced from branches were planted in pots for subsequent experiments. Drought stress was induced by spontaneous evaporation of pots. Leaves were respectively collected as assay samples when the soil water content of pots were 82, 70, 60, 48, 34 and 80% during treatment. Leaves of the 9th, 10th and 11th nods of the stem tips downward from individual plants were taken as one biological replicate samples, and 4 replicates from 4 individual plants were taken each time. The fresh leaves were cut into 5mm × 5mm tissue blocks along the main vein，and then fixed with 70% formaldehyde-acetic-acid-alcohol fixative. After dehydration, each sample were added pure wax and pure transparent agent, and then embedded into wax blocks. They were sliced at 8-10 um thickness by a slicing machine (Mettler Toledo MTT-2010, Mettler Toledo), which were observed on an Olympus biological inverted microscope (OLYMPUSIX73, Olympus) after unfolding, staining and sealing. Each sample was homogenized in an extraction solution (90mM Tris-HCl (pH 7.5), 8% glycerol, 1mM Na2EDTA, 10mM MgCL2, 1mM ascorbic acid and 1.5% polyvinyl pyrrolidone), and centrifuged for 15 min at 12000 rpm and 4°C. The resultant supernatant was used for SOD, APX, POD and CAT activities as well as the levels of soluble proteins and proline. These indexes and the chlorophyll content was determined by ultraviolet spectrophotometry (UV-5100, Metash). The leaf photosynthetic parameter was measured by using an infrared gas analyzer (Li-6400 Portable Photosynthesis System, LI-COR). Samples were homogenized in extraction buffer (8 M urea, 2 mM EDTA, 10 mM DTT, 1% PMSF (Beyotime)), centrifuged, and proteins were acetone-precipitated. The pellet was redissolved in urea-TEAB buffer. After reduction, alkylation, and dilution, proteins were digested with trypsin (1:50, overnight, 37°C; then 1:100, 4 h). Peptides were desalted (Strata X SPE column), fractionated by high-pH reverse-phase HPLC (Waters Bridge Peptide BEH C18 column), and combined into 20 fractions which were desalted (ZipTip C18, Merck Millipore) and dried. LC-MS/MS was performed on an UltiMate 3000 system coupled to an Q Executive HF(Thermo fishier). Peptides were separated on an Acclaim C18 column with an acetonitrile gradient. MS was operated in data-independent mode: full scans (resolution 60,000, m/z 350-1800) were followed by CID fragmentation of the top 20 precursors.MS/MS data were searched against the UniProt Database using SEQUEST in Proteome Discoverer 1.3 . Search parameters included: trypsin digestion, two missed cleavages, 20 ppm precursor tolerance, 0.02 Da fragment tolerance, FDR < 0.1%, fixed modification of carbamidomethylation, and variable modifications of oxidation and N-terminal acetylation.