Abstract
RNA is a key player in the process of gene expression. Whereas fluorescence in situ hybridization allows single mRNA imaging in fixed cells, the MS2-GFP labeling technique enables the observation of mRNA dynamics in living cells. Recently, two genetically engineered mouse models have been developed for the application of the MS2-GFP system in live animals. First, the Actb-MBS mouse was generated by knocking in 24 repeats of the MS2 stem-loop sequence in the 3′ untranslated region of the β-actin gene. Second, the MCP mouse was made to express the NLS-HA-MCP-GFP transgene in all cell types. By crossing Actb-MBS and MCP mice, a double homozygous mouse line, MCP × MBS, was established to visualize endogenous β-actin mRNA labeled with multiple green fluorescent proteins. By imaging hippocampal neurons or brain slices from MCP × MBS mice, the dynamics of mRNA, such as transcription, transport, and localization, can be studied at single mRNA resolution. In this chapter, we explain the basics of MCP × MBS mice and describe methods for utilizing these animals.
| Original language | English (US) |
|---|---|
| Title of host publication | Methods in Enzymology |
| Publisher | Academic Press Inc. |
| Pages | 51-64 |
| Number of pages | 14 |
| DOIs | |
| State | Published - 2016 |
| Externally published | Yes |
Publication series
| Name | Methods in Enzymology |
|---|---|
| Volume | 572 |
| ISSN (Print) | 0076-6879 |
| ISSN (Electronic) | 1557-7988 |
Bibliographical note
Funding Information:This work was supported by the Seoul National University Research Grant in 2015 and Basic Science Research Program through the National Research Foundation of Korea (NRF) funded by the Ministry of Science, ICT & Future Planning (2015R1C1A1A02036674).
Publisher Copyright:
© 2016 Elsevier Inc.
Keywords
- Brain slice
- mRNA
- mRNA localization
- MS2-GFP
- Neuron
- Single-molecule imaging
- Transcription
- Two-photon microscopy
- β-Actin