Affordable Access

deepdyve-link
Publisher Website

Multidimensional quantitative analysis of mRNA expression within intact vertebrate embryos.

Authors
  • Trivedi, Vikas1, 2, 3, 4
  • Choi, Harry M T1
  • Fraser, Scott E2, 3
  • Pierce, Niles A5, 6, 7
  • 1 Division of Biology and Biological Engineering, California Institute of Technology, Pasadena, CA 91125, USA.
  • 2 Translational Imaging Center, University of Southern California, Los Angeles, CA 90089, USA.
  • 3 Molecular and Computational Biology, University of Southern California, Los Angeles, CA 90089, USA.
  • 4 Department of Genetics, University of Cambridge, Cambridge CB2 3EH, UK.
  • 5 Division of Biology and Biological Engineering, California Institute of Technology, Pasadena, CA 91125, USA [email protected]
  • 6 Division of Engineering & Applied Science, California Institute of Technology, Pasadena, CA 91125, USA.
  • 7 Weatherall Institute of Molecular Medicine, University of Oxford, Oxford OX3 9DS, UK.
Type
Published Article
Journal
Development
Publisher
The Company of Biologists
Publication Date
Jan 08, 2018
Volume
145
Issue
1
Identifiers
DOI: 10.1242/dev.156869
PMID: 29311262
Source
Medline
Keywords
License
Unknown

Abstract

For decades, in situ hybridization methods have been essential tools for studies of vertebrate development and disease, as they enable qualitative analyses of mRNA expression in an anatomical context. Quantitative mRNA analyses typically sacrifice the anatomy, relying on embryo microdissection, dissociation, cell sorting and/or homogenization. Here, we eliminate the trade-off between quantitation and anatomical context, using quantitative in situ hybridization chain reaction (qHCR) to perform accurate and precise relative quantitation of mRNA expression with subcellular resolution within whole-mount vertebrate embryos. Gene expression can be queried in two directions: read-out from anatomical space to expression space reveals co-expression relationships in selected regions of the specimen; conversely, read-in from multidimensional expression space to anatomical space reveals those anatomical locations in which selected gene co-expression relationships occur. As we demonstrate by examining gene circuits underlying somitogenesis, quantitative read-out and read-in analyses provide the strengths of flow cytometry expression analyses, but by preserving subcellular anatomical context, they enable bi-directional queries that open a new era for in situ hybridization.

Report this publication

Statistics

Seen <100 times