scholarly journals Whole-Mount RNA In Situ Hybridization and Immunofluorescence of Xenopus Embryos and Tadpoles

2021 ◽  
Vol 2021 (10) ◽  
pp. pdb.prot105635
Author(s):  
Helen Rankin Willsey
Keyword(s):  
In Situ Hybridization ◽  
Final Section ◽  
Stages Of Development ◽  
Xenopus Embryos ◽  
Whole Mount ◽  
Rna In Situ Hybridization ◽  
Endogenous Proteins

A major advantage of experimentation in Xenopus is the ability to query the localization of endogenous proteins and RNAs in situ in the entire animal during all of development. Here I describe three variations of staining to visualize mRNAs and proteins in developing Xenopus embryos and tadpoles. The first section outlines a traditional colorimetric staining for mRNAs that is suitable for all stages of development, and the second extends this protocol for fluorescence-based detection for higher spatial and quantitative resolution. The final section details detection of proteins by immunofluorescence, optimized for tadpole stages but widely applicable to others. Finally, optimization strategies are provided.

Localization of specific mRNAs in Xenopus embryos by whole-mount in situ hybridization

Development ◽  
1990 ◽  
Vol 110 (2) ◽  
pp. 325-330 ◽  
Author(s):  
A. Hemmati-Brivanlou ◽  
D. Frank ◽  
M.E. Bolce ◽  
B.D. Brown ◽  
H.L. Sive ◽  
...  
Keyword(s):  
In Situ Hybridization ◽  
Antisense Rna ◽  
Muscle Actin ◽  
Xenopus Embryos ◽  
Whole Mount ◽  
Alpha Globin ◽  
Specific Mrnas ◽  
Protein Rich ◽  
Hatching Gland

We have adapted a non-radioactive technique to detect localized mRNAs in whole-mount Xenopus embryos. Synthetic antisense RNA transcribed in the presence of digoxygenin-UTP is used as a probe and is detected via an anti-digoxygenin antibody. We show that localized mRNAs can be detected from late gastrula to tadpole stages and that high as well as low abundance RNAs can be detected. The method was tested on muscle actin and alpha-globin RNAs, whose localization has previously been characterized. In addition, we used the method to determine the distribution of XA-1 RNA, an anterior ectoderm-specific RNA, which we show is expressed in the periphery of the cement gland as well as in the region of the hatching gland. The sequence of an XA-1 cDNA predicts a protein rich in proline and histidine.

Whole-Mount In Situ Hybridization and Immunohistochemistry in Xenopus Embryos

2015 ◽  
pp. 151-167 ◽  
Author(s):  
Edgar M. Pera ◽  
Helena Acosta ◽  
Nadège Gouignard ◽  
Maria Climent
Keyword(s):  
In Situ Hybridization ◽  
Xenopus Embryos ◽  
Whole Mount
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