No two clones are alike: characterization of heterologous subpopulations in a transgenic cell line of the model diatom Phaeodactylum tricornutum

Diaz-Garza, A. M., Merindol, N., dos Santos, K. C. G., Lavoie-Marchand, F., Ingalls, B. et Desgagné-Penix, I. (2024). No two clones are alike: characterization of heterologous subpopulations in a transgenic cell line of the model diatom Phaeodactylum tricornutum. Microbial Cell Factories, 23 (1). Article 286. ISSN 1475-2859 DOI 10.1186/s12934-024-02559-y

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Résumé

Abstract

Background:
Conjugation-based episome delivery is a highly efficient method used to transfer DNA into the diatom Phaeodactylum tricornutum, facilitating the production of recombinant proteins and high-value metabolites. However, previous reports have indicated phenotypic heterogeneity among individual cells from clonally propagated exconjugant cell lines, potentially affecting the stability of recombinant protein production in the diatom.

Results:
Here, we characterized the differences between subpopulations with distinct fluorescence intensity phenotypes derived from a single exconjugant colony of P. tricornutum expressing the enhanced green fluorescent protein (eGFP). We analyzed the expression cassette sequence integrity, plasmid copy number, and global gene expression. Our findings reveal that lower copy numbers and the deletion of the expression cassette in part of the population contributed to low transgene expression. Gene co-expression analysis identified a set of genes with similar expression pattern to eGFP including a gene encoding a putative Flp recombinase, which may be related to variations in fluorescence intensity. These genes thus present themselves as potential candidates for increasing recombinant proteins production in P. tricornutum episomal expression system.

Conclusions:
Overall, our study elucidates genetic and transcriptomic differences between distinct subpopulations in a clonally propagated culture, contributes to a better understanding of heterogeneity in diatom expression systems for synthetic biology applications.

Type de document:	Article
Mots-clés libres:	Diatom Heterologous subpopulations Flow cytometry Plasmid copy number Transcriptomics
Date de dépôt:	22 avr. 2025 19:29
Dernière modification:	22 avr. 2025 19:29
Version du document déposé:	Version officielle de l'éditeur
URI:	https://depot-e.uqtr.ca/id/eprint/11866

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