Skip to main content

Lund University Publications

LUND UNIVERSITY LIBRARIES

Ribosome Profiling in the Model Diatom Thalassiosira pseudonana

Pichler, Monica ; Meindl, Andreas ; Romberger, Markus ; Eckes-Shephard, Annemarie LU orcid ; Nyberg-Brodda, Carl Fredrik ; Buhigas, Claudia ; Llaneza-Lago, Sergio ; Lehmann, Gerhard ; Hopes, Amanda and Meister, Gunter , et al. (2023) In Current protocols 3(7).
Abstract

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... (More)

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.

(Less)
Please use this url to cite or link to this publication:
author
; ; ; ; ; ; ; ; and , et al. (More)
; ; ; ; ; ; ; ; ; ; and (Less)
organization
publishing date
type
Contribution to journal
publication status
published
subject
keywords
diatoms, high-throughput sequencing, ribosome profiling, RNA, Thalassiosira pseudonana, translation
in
Current protocols
volume
3
issue
7
article number
e843
publisher
John Wiley & Sons Inc.
external identifiers
  • pmid:37439534
  • scopus:85164540656
ISSN
2691-1299
DOI
10.1002/cpz1.843
language
English
LU publication?
yes
additional info
Publisher Copyright: © 2023 The Authors. Current Protocols published by Wiley Periodicals LLC.
id
3d404ff7-1808-4c4c-b02d-73b59213537d
date added to LUP
2023-07-26 11:00:41
date last changed
2024-04-19 23:58:10
@article{3d404ff7-1808-4c4c-b02d-73b59213537d,
  abstract     = {{<p>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.</p>}},
  author       = {{Pichler, Monica and Meindl, Andreas and Romberger, Markus and Eckes-Shephard, Annemarie and Nyberg-Brodda, Carl Fredrik and Buhigas, Claudia and Llaneza-Lago, Sergio and Lehmann, Gerhard and Hopes, Amanda and Meister, Gunter and Medenbach, Jan and Mock, Thomas}},
  issn         = {{2691-1299}},
  keywords     = {{diatoms; high-throughput sequencing; ribosome profiling; RNA; Thalassiosira pseudonana; translation}},
  language     = {{eng}},
  month        = {{07}},
  number       = {{7}},
  publisher    = {{John Wiley & Sons Inc.}},
  series       = {{Current protocols}},
  title        = {{Ribosome Profiling in the Model Diatom Thalassiosira pseudonana}},
  url          = {{http://dx.doi.org/10.1002/cpz1.843}},
  doi          = {{10.1002/cpz1.843}},
  volume       = {{3}},
  year         = {{2023}},
}