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Annemarie Eckes-Shephard

Annemarie Eckes-Shephard

Researcher

Annemarie Eckes-Shephard

Ribosome Profiling in the Model Diatom Thalassiosira pseudonana

Author

  • Monica Pichler
  • Andreas Meindl
  • Markus Romberger
  • Annemarie Eckes-Shephard
  • Carl Fredrik Nyberg-Brodda
  • Claudia Buhigas
  • Sergio Llaneza-Lago
  • Gerhard Lehmann
  • Amanda Hopes
  • Gunter Meister
  • Jan Medenbach
  • Thomas Mock
Show all

Summary, in English

Diatoms are an important group of eukaryotic microalgae, which play key roles in marine biochemical cycling and possess significant biotechnological potential. Despite the importance of diatoms, their regulatory mechanisms of protein synthesis at the translational level remain largely unexplored. Here, we describe the detailed development of a ribosome profiling protocol to study translation in the model diatom Thalassiosira pseudonana, which can easily be adopted for other diatom species. To isolate and sequence ribosome-protected mRNA, total RNA was digested, and the ribosome-protected fragments were obtained by a combination of sucrose-cushion ultracentrifugation and polyacrylamide gel electrophoresis for size selection. To minimize rRNA contamination, a subtractive hybridization step using biotinylated oligos was employed. Subsequently, fragments were converted into sequencing libraries, enabling the global quantification and analysis of changes in protein synthesis in diatoms. The development of this novel ribosome profiling protocol represents a major expansion of the molecular toolbox available for diatoms and therefore has the potential to advance our understanding of the translational regulation in this important group of phytoplankton.

Department/s

  • Dept of Physical Geography and Ecosystem Science

Publishing year

2023-07-01

Language

English

Publication/Series

Current protocols

Volume

3

Issue

7

Document type

Journal article

Publisher

John Wiley & Sons Inc.

Topic

  • Biochemistry and Molecular Biology

Keywords

  • diatoms
  • high-throughput sequencing
  • ribosome profiling
  • RNA
  • Thalassiosira pseudonana
  • translation

Status

Published

ISBN/ISSN/Other

  • ISSN: 2691-1299