Project PXD000625

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Summary

Title

Hepatocyte-specific BIRC5 (Survivin)-knockout-mouse model, Subcellular fractionation

Description

Experimental design For the label-free proteome analysis 17 mice were used composed of 5 individual wildtype mice for each condition before (WTprePHE) and after hepatectomy (WTpostPHE). For the pre-hepatectomy condition four BIRC5-knockout mice (KOprePHE) were analyzed and three individuals after partial hepatectomy (KOpostPHE). For the LC-MS/MS analysis the samples were pre-fractionated by subcellular protein fractionatio. All samples corresponding to a subcellular fraction were analyzed in an individual label-free study. Subcellular protein fractionation The subcellular fractions were generated using the Subcellular Protein Fractionation Kit for Tissue (Thermo Scientific, Bremen, Germany). Sample preparation and tryptic digestion 5ug of protein were loaded on a 18% Tris-glycine acrylamide gel (Anamed Electrophorese, Grosz-Bieberau, Germany). The samples were allowed to run slightly into the gel (15 min, 100 V) and form a single band of approximately 3mm height. The bands were stained with Coomassie, manually cut from the gel and digested with trypsin (Serva Electrophoresis, Heidelberg, Germany) in 10 mM ammonium bicarbonate buffer (pH 7.8) overnight at 37 degrees C. LC-MS/MS Analysis The RPLC-MS/MS analysis was performed using an Ultimate 3000 RSLCnano system (Thermo Scientific, Bremen, Germany) online coupled to an Orbitrap Elite mass spectrometer (Thermo Scientific, Bremen, Germany). The injection volume was 15 ul of the sample, representing an amount of 250 ng peptides.The MS was operated in a data-dependent mode. Full scan MS spectra were acquired at a resolution of 60,000 in the Orbitrap analyzer, while tandem mass spectra of the twenty most abundant peaks were measured in the linear ion trap after peptide fragmentation by collision-induced dissociation (CID).

Sample Processing Protocol

See details in reference(s) : 24818710

Data Processing Protocol

See details in reference(s) : 24818710

Contact

Thilo Bracht, Clinical Proteomics

Submission Date

17/12/2013

Publication Date

22/05/2014

Tissue

liver

Quantification

Not available

Experiment Type

Bottom-up proteomics

Assay count

68

Publication

    Bracht T, Hagemann S, Loscha M, Megger DA, Padden J, Eisenacher M, Kuhlmann K, Meyer HE, Baba HA, Sitek B. Proteome analysis of a hepatocyte-specific BIRC5(Survivin)-knockout mouse model during liver regeneration. J Proteome Res. 2014 May 12 PubMed: 24818710

Assay

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Showing 1 - 10 of 68 results
# Accession Title Proteins Peptides Unique Peptides Spectra Identified Spectra View in Reactome
1 33250 Analysis of a hepatocyte-specific BIRC5 (Survivin)-knockout mouse model during liver regeneration induced by partial hepatectomy - Experiment 1 1239 8390 3510 25213 7266
2 33251 Analysis of a hepatocyte-specific BIRC5 (Survivin)-knockout mouse model during liver regeneration induced by partial hepatectomy - Experiment 2 1902 10820 6773 24406 9645
3 33252 Analysis of a hepatocyte-specific BIRC5 (Survivin)-knockout mouse model during liver regeneration induced by partial hepatectomy - Experiment 3 1943 10295 6900 26855 9231
4 33253 Analysis of a hepatocyte-specific BIRC5 (Survivin)-knockout mouse model during liver regeneration induced by partial hepatectomy - Experiment 4 1921 10867 7157 26078 9716
5 33254 Analysis of a hepatocyte-specific BIRC5 (Survivin)-knockout mouse model during liver regeneration induced by partial hepatectomy - Experiment 5 1710 8711 5758 24332 7703
6 33255 Analysis of a hepatocyte-specific BIRC5 (Survivin)-knockout mouse model during liver regeneration induced by partial hepatectomy - Experiment 6 1985 10082 6614 26355 8982
7 33256 Analysis of a hepatocyte-specific BIRC5 (Survivin)-knockout mouse model during liver regeneration induced by partial hepatectomy - Experiment 7 1361 6214 3956 21934 5519
8 33257 Analysis of a hepatocyte-specific BIRC5 (Survivin)-knockout mouse model during liver regeneration induced by partial hepatectomy - Experiment 8 1991 10476 7011 25736 9324
9 33258 Analysis of a hepatocyte-specific BIRC5 (Survivin)-knockout mouse model during liver regeneration induced by partial hepatectomy - Experiment 9 1866 10711 6906 27182 9602
10 33259 Analysis of a hepatocyte-specific BIRC5 (Survivin)-knockout mouse model during liver regeneration induced by partial hepatectomy - Experiment 10 2044 11173 7362 25747 9892