Isolation of Protein Complexes Involved in Mitosis and Cytokinesis from Drosophila Cultured Cells
The identification of all the individual components that constitute the plethora of complexes in each cell type represents perhaps the most exciting challenge of postgenomic biology. This is particularly important in the study of events such as mitosis and cytokinesis, in which rapid and precise protein–protein interactions regulate both the direction and accuracy of these intricate processes. Here we describe an experimental strategy to isolate protein complexes involved in mitosis and cytokinesis in cultured Drosophila cells. This method involves the tagging of the bait protein with two IgG binding domains of Protein A and the isolation of the tagged bait along with its interacting partners by a single affinity purification step. These isolated complexes can then be analysed by several methods including mass spectrometry and Western blotting. Although this method has proven very successful in isolating mitotic and cytokinetic complexes, it can also be used to characterise protein complexes involved in many other cellular processes.
- Using 3D Culture to Investigate the Role of Mechanical Signaling in Keratinocyte Stem Cells
- Measurements of Peptide and Nonpeptide Secretory Products from Adipocytes
- 大鼠血红蛋白(Hb)酶联免疫分析(ELISA)
- DNA Microarrays to Analyze Gene Expression in Animals with Altered Transporter Expression
- The Application of Randomly Amplified DNA Analysis in the Molecular Epidemiology of Microorganisms
- Cell Viability Analysis Using Trypan Blue: Manual and Automated Methods
- Analysis of Tissue-Specific Gene Expression Using Laser Capture Microdissection
- Carcinogenesis Studies in Mice with Genetically Engineered Alterations in Polyamine Metabolism
- Nonviral Gene Delivery in Neural Progenitors Derived from Human Pluripotent Stem Cells
- 体外细胞的原代培养、传代培养、冻存和复苏实验方法步骤