Cell type-specific translational profiling in the Xenopus laevis retina

F. L. Watson, E. A. Mills, X. Wang, C. Guo, D. F. Chen, Nicholas Marsh-Armstrong

Research output: Contribution to journalArticle

18 Scopus citations

Abstract

Background: Translating Ribosome Affinity Purification (TRAP), a method recently developed to generate cell type-specific translational profiles, relies on creating transgenic lines of animals in which a tagged ribosomal protein is placed under regulatory control of a cell type-specific promoter. An antibody is then used to affinity purify the tagged ribosomes so that cell type-specific mRNAs can be isolated from whole tissue lysates. Results: Here, cell type-specific transgenic lines were generated to enable TRAP studies for retinal ganglion cells and rod photoreceptors in the Xenopus laevis retina. Using real time quantitative PCR for assessing expression levels of cell type-specific mRNAs, the TRAP method was shown to selectively isolate mRNAs expressed in the targeted cell and was efficient at purifying mRNAs expressed at both high and low levels. Statistical measures used to distinguish cell type-specific RNAs from low level background and non-specific RNAs showed TRAP to be highly effective in Xenopus. Conclusions: TRAP can be used to purify mRNAs expressed in rod photoreceptors and retinal ganglion cells in X. laevis. The generated transgenic lines will enable numerous studies into the development, disease, and injury of the X. laevis retina.

Original languageEnglish (US)
Pages (from-to)1960-1972
Number of pages13
JournalDevelopmental Dynamics
Volume241
Issue number12
DOIs
StatePublished - Dec 1 2012
Externally publishedYes

Keywords

  • Cell type-specific transgenic lines
  • Retina
  • Retinal ganglion cells
  • Rod photoreceptor cells
  • Translating ribosome affinity purification
  • TRAP
  • Xenopus laevis

ASJC Scopus subject areas

  • Developmental Biology

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