Minimizing Clonal Variation during Mammalian Cell Line Engineering for Improved Systems Biology Data Generation

Lise Marie Grav, Daria Sergeeva, Jae Seong Lee, Igor Marín de Mas, Nathan E. Lewis, Mikael Rørdam Andersen, Lars Keld Nielsen, Gyun Min Lee, Helene Faustrup Kildegaard*

*Corresponding author for this work

Research output: Contribution to journalJournal articleResearchpeer-review

Abstract

Mammalian cells are widely used to express genes for basic biology studies and biopharmaceuticals. Current methods for generation of engineered cell lines introduce high genomic and phenotypic diversity, which hamper studies of gene functions and discovery of novel cellular mechanisms. Here, we minimized clonal variation by integrating a landing pad for recombinase-mediated cassette exchange site-specifically into the genome of CHO cells using CRISPR and generated subclones expressing four different recombinant proteins. The subclones showed low clonal variation with high consistency in growth, transgene transcript levels and global transcriptional response to recombinant protein expression, enabling improved studies of the impact of transgenes on the host transcriptome. Little variation over time in subclone phenotypes and transcriptomes was observed when controlling environmental culture conditions. The platform enables robust comparative studies of genome engineered CHO cell lines and can be applied to other mammalian cells for diverse biological, biomedical and biotechnological applications.
Original languageEnglish
JournalA C S Synthetic Biology
Volume7
Issue number9
Pages (from-to)2148-2159
ISSN2161-5063
DOIs
Publication statusPublished - 2018

Keywords

  • Clonal variation
  • CRISPR/Cas9
  • Mammalian cells
  • Recombinase-mediated cassette exchange
  • Targeted integration
  • Transcriptome

Cite this

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title = "Minimizing Clonal Variation during Mammalian Cell Line Engineering for Improved Systems Biology Data Generation",
abstract = "Mammalian cells are widely used to express genes for basic biology studies and biopharmaceuticals. Current methods for generation of engineered cell lines introduce high genomic and phenotypic diversity, which hamper studies of gene functions and discovery of novel cellular mechanisms. Here, we minimized clonal variation by integrating a landing pad for recombinase-mediated cassette exchange site-specifically into the genome of CHO cells using CRISPR and generated subclones expressing four different recombinant proteins. The subclones showed low clonal variation with high consistency in growth, transgene transcript levels and global transcriptional response to recombinant protein expression, enabling improved studies of the impact of transgenes on the host transcriptome. Little variation over time in subclone phenotypes and transcriptomes was observed when controlling environmental culture conditions. The platform enables robust comparative studies of genome engineered CHO cell lines and can be applied to other mammalian cells for diverse biological, biomedical and biotechnological applications.",
keywords = "Clonal variation, CRISPR/Cas9, Mammalian cells, Recombinase-mediated cassette exchange, Targeted integration, Transcriptome",
author = "Grav, {Lise Marie} and Daria Sergeeva and Lee, {Jae Seong} and {Mar{\'i}n de Mas}, Igor and Lewis, {Nathan E.} and Andersen, {Mikael R{\o}rdam} and Nielsen, {Lars Keld} and {Min Lee}, Gyun and Kildegaard, {Helene Faustrup}",
year = "2018",
doi = "10.1021/acssynbio.8b00140",
language = "English",
volume = "7",
pages = "2148--2159",
journal = "A C S Synthetic Biology",
issn = "2161-5063",
publisher = "American Chemical Society",
number = "9",

}

Minimizing Clonal Variation during Mammalian Cell Line Engineering for Improved Systems Biology Data Generation. / Grav, Lise Marie; Sergeeva, Daria; Lee, Jae Seong; Marín de Mas, Igor; Lewis, Nathan E.; Andersen, Mikael Rørdam; Nielsen, Lars Keld; Min Lee, Gyun; Kildegaard, Helene Faustrup.

In: A C S Synthetic Biology, Vol. 7, No. 9, 2018, p. 2148-2159.

Research output: Contribution to journalJournal articleResearchpeer-review

TY - JOUR

T1 - Minimizing Clonal Variation during Mammalian Cell Line Engineering for Improved Systems Biology Data Generation

AU - Grav, Lise Marie

AU - Sergeeva, Daria

AU - Lee, Jae Seong

AU - Marín de Mas, Igor

AU - Lewis, Nathan E.

AU - Andersen, Mikael Rørdam

AU - Nielsen, Lars Keld

AU - Min Lee, Gyun

AU - Kildegaard, Helene Faustrup

PY - 2018

Y1 - 2018

N2 - Mammalian cells are widely used to express genes for basic biology studies and biopharmaceuticals. Current methods for generation of engineered cell lines introduce high genomic and phenotypic diversity, which hamper studies of gene functions and discovery of novel cellular mechanisms. Here, we minimized clonal variation by integrating a landing pad for recombinase-mediated cassette exchange site-specifically into the genome of CHO cells using CRISPR and generated subclones expressing four different recombinant proteins. The subclones showed low clonal variation with high consistency in growth, transgene transcript levels and global transcriptional response to recombinant protein expression, enabling improved studies of the impact of transgenes on the host transcriptome. Little variation over time in subclone phenotypes and transcriptomes was observed when controlling environmental culture conditions. The platform enables robust comparative studies of genome engineered CHO cell lines and can be applied to other mammalian cells for diverse biological, biomedical and biotechnological applications.

AB - Mammalian cells are widely used to express genes for basic biology studies and biopharmaceuticals. Current methods for generation of engineered cell lines introduce high genomic and phenotypic diversity, which hamper studies of gene functions and discovery of novel cellular mechanisms. Here, we minimized clonal variation by integrating a landing pad for recombinase-mediated cassette exchange site-specifically into the genome of CHO cells using CRISPR and generated subclones expressing four different recombinant proteins. The subclones showed low clonal variation with high consistency in growth, transgene transcript levels and global transcriptional response to recombinant protein expression, enabling improved studies of the impact of transgenes on the host transcriptome. Little variation over time in subclone phenotypes and transcriptomes was observed when controlling environmental culture conditions. The platform enables robust comparative studies of genome engineered CHO cell lines and can be applied to other mammalian cells for diverse biological, biomedical and biotechnological applications.

KW - Clonal variation

KW - CRISPR/Cas9

KW - Mammalian cells

KW - Recombinase-mediated cassette exchange

KW - Targeted integration

KW - Transcriptome

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SN - 2161-5063

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