Preparation Methods of Human Metaphase Chromosomes for their Proteome Analysis.
Chromosomes are supermolecules that contain most of the DNA within a cell and are visible under optical and electron microscopes. Although they were observed at the earliest stage of genetics, their fundamental structure is not yet understood. The reasons for this are debated among researchers; however, it is clear that the accumulation of metaphase chromosomes for their biochemical analysis has been a significant challenge. In this chapter, a method is described for accumulating and preparing human metaphase chromosomes in sufficient amount to perform their proteome analysis. Preparation and separation methods of chromosome proteins are described, followed by a protocol for their identification by mass spectrometry.
Fukui
Kiichi
K
Department of Biotechnology, Graduate School of Engineering, Osaka University, Osaka, Japan.
Takata
Hideaki
H
Uchiyama
Susumu
S
D002460:Cell Line; D002462:Cell Membrane / Q000648:ultrastructure; D002868:Chromosomal Proteins, Non-Histone / Q000737:chemistry / Q000302:isolation & purification; D002877:Chromosomes, Human / Q000737:chemistry* / Q000648:ultrastructure*; D003703:Demecolcine; D015180:Electrophoresis, Gel, Two-Dimensional / Q000379:methods; D004591:Electrophoresis, Polyacrylamide Gel / Q000379:methods; D006367:HeLa Cells; D006801:Humans; D007202:Indicators and Reagents; D013058:Mass Spectrometry / Q000379:methods; D009687:Nuclear Proteins / Q000737:chemistry* / Q000302:isolation & purification*; D020543:Proteome; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization / Q000379:methods
D002868:Chromosomal Proteins, Non-Histone; D007202:Indicators and Reagents; D009687:Nuclear Proteins; D020543:Proteome; D003703:Demecolcine