DICP OpenIR
Subject Area物理化学
Integration of Cell Lysis, Protein Extraction, and Digestion into One Step for Ultrafast Sample Preparation for Phosphoproteome Analysis
Liu, Fangjie1,2; Ye, Mingliang1; Pan, Yanbo1,2; Zhang, Yi1,2; Bian, Yangyang1,2; Sun, Zhen1,2; Zhu, Jun1,2; Cheng, Kai1,2; Zou, Hanfa1; Zou HF(邹汉法); Ye ML(叶明亮)
Source PublicationANALYTICAL CHEMISTRY
2014-07-15
ISSN0003-2700
DOI10.1021/ac5002146
Volume86Issue:14Pages:6786-6791
Indexed BySCI
Cooperation Status
SubtypeArticle
Department18
Funding Project1809
Contribution Rank待补充
WOS HeadingsScience & Technology ; Physical Sciences
Funding Organization1,1 ; 1,1 ; 1,1 ; 1,1
WOS SubjectChemistry, Analytical
WOS Research AreaChemistry
WOS KeywordIMMOBILIZED TRYPSIN DIGESTION ; INTENSITY FOCUSED ULTRASOUND ; PROTEOMICS ; CHROMATOGRAPHY ; IMPACT ; MASS
AbstractConventional sample preparation protocols for phosphoproteome analysis require multiple time-consuming and labor-intensive steps, including cell lysis, protein extraction, protein digestion, and phosphopeptide enrichment. In this study, we found that the presence of a large amount of trypsin in the sample did not interfere with phosphopeptide enrichment and subsequent LC-MS/MS analysis. Taking advantage of fast digestion achieved with high trypsin-to-protein ratio, we developed a novel concurrent lysis-digestion method for phosphoproteome analysis. In this method, the harvested cells were first placed in a lysis buffer containing a huge amount of trypsin. After ultrasonication, the cells were lysed and the proteins were efficiently digested into peptides within one step. Thereafter, tryptic digest was subjected to phosphopeptide enrichment, in which unphosphorylated peptides, trypsin, and other components incompatible with LC-MS/MS analysis were removed. Compared with conventional methods, better phosphoproteome coverage was achieved in this new one-step method. Because protein solubilization and cell lysis were facilitated by fast protein digestion, the complete transformation of cell pellets into the peptide mixture could be finished within 25 min, while it would take at least 16 h for conventional methods. Hence, our method, which integrated cell lysis, protein extraction, and protein digestion into one step, is rapid and convenient. It is expected to have broad applications in phosphoproteomics analysis.
Language英语
Funding Organization1,1 ; 1,1 ; 1,1 ; 1,1
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WOS IDWOS:000339227400010
Citation statistics
Cited Times:12[WOS]   [WOS Record]     [Related Records in WOS]
Document Type期刊论文
Identifierhttp://cas-ir.dicp.ac.cn/handle/321008/119725
Collection中国科学院大连化学物理研究所
Corresponding AuthorZou HF(邹汉法); Ye ML(叶明亮)
Affiliation1.Chinese Acad Sci, Dalian Inst Chem Phys, Key Lab Separat Sci Analyt Chem, Dalian 116023, Liaoning, Peoples R China
2.Chinese Acad Sci, Grad Sch, Beijing 100049, Peoples R China
Recommended Citation
GB/T 7714
Liu, Fangjie,Ye, Mingliang,Pan, Yanbo,et al. Integration of Cell Lysis, Protein Extraction, and Digestion into One Step for Ultrafast Sample Preparation for Phosphoproteome Analysis[J]. ANALYTICAL CHEMISTRY,2014,86(14):6786-6791.
APA Liu, Fangjie.,Ye, Mingliang.,Pan, Yanbo.,Zhang, Yi.,Bian, Yangyang.,...&叶明亮.(2014).Integration of Cell Lysis, Protein Extraction, and Digestion into One Step for Ultrafast Sample Preparation for Phosphoproteome Analysis.ANALYTICAL CHEMISTRY,86(14),6786-6791.
MLA Liu, Fangjie,et al."Integration of Cell Lysis, Protein Extraction, and Digestion into One Step for Ultrafast Sample Preparation for Phosphoproteome Analysis".ANALYTICAL CHEMISTRY 86.14(2014):6786-6791.
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