Large-scale expression screening by automated whole-mount in situ hybridization

Rebecca Quiring, Beate Wittbrodt, Thorsten Henrich, Mirana Ramialison, Carola Burgtorf, Hans Lehrach, Joachim Wittbrodt

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38 Citations (Scopus)

Abstract

Gene expression profiling is an important component of functional genomics. We present a time and cost efficient high-throughput whole-mount in situ technique to perform a large-scale gene expression analysis in medaka fish (Oryzias latipes) embryos. Medaka is a model system ideally suited for the study of molecular genetics of vertebrate development. Random cDNA clones from an arrayed stage 20 medaka plasmid library were analyzed by whole-mount in situ hybridization on embryos of three representative stages of medaka development. cDNA inserts were colony PCR amplified in a 384-format. The PCR products were used to generate over 2000 antisense RNA digoxigenin probes in a high-throughput process. Whole-mount in situ hybridization was carried out in a robot and a broad range of expression patterns was observed. Partial cDNA sequences and expression patterns were documented with BLAST results, cluster analysis, images and descriptions, respectively; collectively this information was entered into a web-based database, 'MEPD' (http://www.embl-heidelberg.de/mepd/), that is publicly accessible.

Original languageEnglish
Pages (from-to)971-976
Number of pages6
JournalMechanisms of Development
Volume121
Issue number7-8
DOIs
Publication statusPublished - Jul 2004
Externally publishedYes

Keywords

  • Automated
  • Database
  • Embryogenesis
  • Expressed Sequence Tags
  • Expression profile
  • Gene expression
  • High-throughput
  • Organogenesis
  • Oryzias latipes
  • Partial cDNA sequencing
  • Screening
  • Whole-mount in situ hybridization

Cite this

Quiring, R., Wittbrodt, B., Henrich, T., Ramialison, M., Burgtorf, C., Lehrach, H., & Wittbrodt, J. (2004). Large-scale expression screening by automated whole-mount in situ hybridization. Mechanisms of Development, 121(7-8), 971-976. https://doi.org/10.1016/j.mod.2004.03.031