Project PXD006833

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Biomedical Dataset
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Summary

Title

Human bladder,colon,kidney,liver cancer LC MS/MS

Description

We propose that enrichment of low-abundance proteins benefits MPs finding. ProteoMinerTM is an equalizing technique by reducing high-abundance proteins and enriching low-abundance proteins in biological liquids. With triton X-100/TBS buffer extraction, ProteoMinerTM enrichment and peptide fractionation, 25 MPs with 73 unique peptides were identified from four human tissues, including 10 membrane proteins and 6 nucleus proteins. Then 18 MPs were confirmed with at least 2 unique peptide identification and 6 MPs were confirmed with 1 unique peptide identification by PRM assay. Hence these results demonstrated ProteoMinerTM as a powerful means in discovery of MPs.

Sample Processing Protocol

Tissues were ground in a tissue lyser and further lysed by probe sonication in the extraction buffer of PBS (pH 7.5), HEPES (20 mM, pH 8.0), NH4HCO3 (50 mM, pH 8.0) or triton buffer (1% TritonX-100, 20 mM Tris, 150 mM NaCl, pH 7.4) with protease inhibitor cocktail at 0-4 �? Bradford assay was adopted to measure the concentrations of extracted proteins with bovine serum albumin as a standard. Lower abundance proteins were enriched with ProteoMinerTM Protein Enrichment Kits according to manufacturer’s protocol. In brief, the columns were conditioned by the extraction buffer and then the extracted tissue proteins were loaded onto them under gravity speed. The nonspecific binding proteins were washed out by the same extraction buffer, and then the specific binding proteins were eluted by the elution buffer provided in the kits. The amount of input proteins and the volume of ProteoMinerTM beads used were calculated according to the kit manual suggestions. The effects of higher abundance protein depletion were evaluated by protein recovery ratio and the images of SDS-PAGE gels to show the protein pattern changes with ProteoMinerTM treatment.

Data Processing Protocol

Acquired MS data were converted to MGF files by Proteome Discoverer 1.4 (Thermo Scientific, Waltham, MA) and the exported MGF files were searched using Mascot version2.3.02 against the Swiss-prot human database ( released on 2017_04 with 20183 protein sequences). The false discovery rate (FDR) was set to less than 1% at both PSM and protein level. Trypsin was selected as the specific enzyme with a maximum of two missed cleavages permitted per peptide. Parameters included fixed modification: Carbamidomethylation (C); variable modification: Oxidation (M), Deamidatioin (N, Q). Data were searched with a peptide mass tolerance of 20ppm and a fragment mass tolerance of 0.05Da.

Contact

He Yanbin, +86 18565610792
Siqi Liu, Professor and Director Center of Proteomic Analysis BGI-Shenzhen ( lab head )

Submission Date

29/06/2017

Publication Date

02/10/2017

Tissue

liver

Instrument

Q Exactive HF

Software

Not available

Experiment Type

Shotgun proteomics

Assay count

15

Publication

    Li S, He Y, Lin Z, Xu S, Zhou R, Liang F, Wang J, Yang H, Liu S, Ren Y. Digging more missing proteins using an enrichment approach with ProteoMiner™. J Proteome Res. 2017 PubMed: 28960076

Assay

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Page size 10
Showing 1 - 10 of 15 results
# Accession Title Proteins Peptides Unique Peptides Spectra Identified Spectra View in Reactome
1 79191 liver_1.dat-pride.xml 12701 247055 49364 897042 0
2 79190 liver_2.dat-pride.xml 12045 227554 45571 848993 0
3 79193 liver_1_2.dat-pride.xml 14778 494243 58554 1746035 0
4 79192 liver_3.dat-pride.xml 12716 245143 53909 899748 0
5 79195 liver_4.dat-pride.xml 12431 240598 52166 861682 0
6 79194 liver_3_4.dat-pride.xml 15033 507841 65837 1761430 0
7 79197 colon_T_1.dat-pride.xml 11488 180447 43720 784050 0
8 79196 colon_T_2.dat-pride.xml 11351 175539 43278 775073 0
9 79188 colon_T_1_2.dat-pride.xml 12910 367344 50494 1559123 0
10 79199 kidney_1.dat-pride.xml 12997 339178 78662 857784 0