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Two-dimensional low-flow liquid-chromatography setup for shotgun proteomics

ABSTRACT: Development of low-flow liquid-chromatography setup by combining hydrophylic interaction liquid chromatography (HILIC) and reversed phase liquid chromatography (RPLC) and coupled with a high resolution mass spectrometer.

INSTRUMENT(S): Q Exactive

ORGANISM(S): Homo Sapiens (human) Saccharomyces Cerevisiae (baker's Yeast)

TISSUE(S): Lung, Kidney

SUBMITTER: Liana Roca

LAB HEAD: Peter J Schoenmakers

PROVIDER: PXD022330 | Pride | 2021-03-29

REPOSITORIES: Pride

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			Action	DRS
	20200116_RP75_15cm_1000ng_FF5mg_120min_1.msf	Msf
	20200116_RP75_15cm_1000ng_FF5mg_120min_1.mzML	Mzml
	20200116_RP75_15cm_1000ng_FF5mg_120min_1.mzid.gz	Mzid
	20200116_RP75_15cm_1000ng_FF5mg_120min_1.raw	Raw
	20200116_RP75_15cm_1000ng_HCT_120min_1.mzML	Mzml

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Project description:The advent of high-throughput measurements of gene expression and bioinformatics analysis methods offers new ways to study gene expression patterns. The primary goal of this study was to determine the time sequence for gene expression in a bone subjected to mechanical loading, during key periods of the bone formation process, including expression of matrix-related genes, the appearance of active osteoblasts, and bone desensitization. A standard model for bone loading was employed in which the right forelimb was loaded axially for three minutes per day, while the left forearm served as a non-loaded, contralateral control. We evaluated loading-induced gene expression over a time course of 4 hours to 32 days after the first loading session. Six distinct, time-dependent patterns of gene expression were identified over the time course and categorized into three primary clusters: genes upregulated early in the time course, genes upregulated during matrix formation, and genes downregulated during matrix formation. Genes were then grouped based on function and/or signaling pathways. Many gene groups known to be important in loading-induced bone formation were identified within the clusters, including AP-1-related genes in the early response cluster, matrix-related genes in the upregulated gene clusters, and Wnt/?-catenin signaling pathway inhibitors in the downregulated gene clusters. Several novel gene groups were identified as well, including chemokine-related genes which were upregulated early but downregulated later in the time course, solute carrier genes which were both up- and downregulated, and muscle-related genes which were primarily downregulated. Time Course with 11 time points, each plus & minus mechanical stimulation with 5 replicates per experimental group (except 12d group which has 4 replicates). Daily mechanical loading was applied to the forearm (24 hours between loading sessions), and ulnae were sampled at indicated time points (4h, 12h, 1d, 2d, 4d, 6d, 8d, 12d, 16d, 24d, or 32d).

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Two-dimensional low-flow liquid-chromatography setup for shotgun proteomics

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