Chapter title |
Analysis of protein dynamics with tandem fluorescent protein timers.
|
---|---|
Chapter number | 13 |
Book title |
Exocytosis and Endocytosis
|
Published in |
Methods in molecular biology, January 2014
|
DOI | 10.1007/978-1-4939-0944-5_13 |
Pubmed ID | |
Book ISBNs |
978-1-4939-0943-8, 978-1-4939-0944-5
|
Authors |
Anton Khmelinskii, Michael Knop, Khmelinskii, Anton, Knop, Michael |
Abstract |
Fluorescent timers (FTs) are fluorescent proteins that change color with time. FTs can be used as tags to follow protein dynamics in living cells. Recently we described a novel class of FTs called tandem fluorescent protein timers (tFTs). Each tFT is a tandem fusion of two different conventional fluorescent proteins having distinct kinetics of fluorophore maturation. tFTs suitable for studying protein dynamics on different scales can be generated from a broad range of commonly used fluorescent proteins. Here we describe how to establish new tFTs and consider potential pitfalls. We detail a protocol for quantitative fluorescence microscopy imaging and analysis of intracellular protein dynamics with tFTs in the budding yeast Saccharomyces cerevisiae. |
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