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An indelible lineage marker for Xenopus using a mutated green fluorescent protein

Zernicka-Goetz, M. and Pines, J. and Ryan, K. and Siemering, K. R. and Haseloff, J. and Evans, M. J. and Gurdon, J. B. (1996) An indelible lineage marker for Xenopus using a mutated green fluorescent protein. Development, 122 (12). pp. 3719-3724. ISSN 0950-1991. http://resolver.caltech.edu/CaltechAUTHORS:20190422-101712629

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Abstract

We describe the use of a DNA construct (named GFP.RN3) encoding green fluorescent protein as a lineage marker for Xenopus embryos. This offers the following advantages over other lineage markers so far used in Xenopus. When injected as synthetic mRNA, its protein emits intense fluorescence in living embryos. It is non-toxic, and the fluorescence does not bleach when viewed under 480 nm light. It is surprisingly stable, being strongly visible up to the feeding tadpole stage (5 days), and in some tissues for several weeks after mRNA injection. We also describe a construct that encodes a blue fluorescent protein. We exemplify the use of this GFP.RN3 construct for marking the lineage of individual blastomeres at the 32- to 64-cell stage, and as a marker for single transplanted blastula cells. Both procedures have revealed that the descendants of one embryonic cell can contribute single muscle cells to nearly all segmental myotomes rather than predominantly to any one myotome. An independent aim of our work has been to follow the fate of cells in which an early regulatory gene has been temporarily overexpressed. For this purpose, we co-injected GFP.RN3 mRNA and mRNA for the early Xenopus gene Eomes, and found that a high concentration of Eomes results in ectopic muscle gene activation in only the injected cells. This marker may therefore be of general value in providing long term identification of those cells in which an early gene with ephemeral expression has been overexpressed.


Item Type:Article
Related URLs:
URLURL TypeDescription
http://dev.biologists.org/content/122/12/3719.longPublisherArticle
ORCID:
AuthorORCID
Zernicka-Goetz, M.0000-0002-7004-2471
Pines, J.0000-0002-5227-6004
Additional Information:© 1996 The Company of Biologists. (Accepted 18 September 1996) We thank F. Stennard and A. Mitchell for their help with some technical procedures, and A. Brand and C. Davidson for pointing out the presence of the H25Y mutation in GFP.RN3. M. Z.-G. thanks EMBO for a Long Term Research Fellowship. This work was supported by a Cancer Research Campaign programme grant SP 2184/0101 to J. B. Gurdon.
Funders:
Funding AgencyGrant Number
European Molecular Biology Organization (EMBO)UNSPECIFIED
Cancer Research CampaignSP 2184/0101
Subject Keywords:Xenopus, green fluorescent protein, marker, Eomes, cell lineage, GFP.RN3
Record Number:CaltechAUTHORS:20190422-101712629
Persistent URL:http://resolver.caltech.edu/CaltechAUTHORS:20190422-101712629
Official Citation:An indelible lineage marker for Xenopus using a mutated green fluorescent protein M. Zernicka-Goetz, J. Pines, K. Ryan, K.R. Siemering, J. Haseloff, M.J. Evans, J.B. Gurdon Development 1996 122: 3719-3724
Usage Policy:No commercial reproduction, distribution, display or performance rights in this work are provided.
ID Code:94848
Collection:CaltechAUTHORS
Deposited By: George Porter
Deposited On:23 Apr 2019 16:52
Last Modified:23 Apr 2019 16:52

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