ArticleBMC biotechnology2016
Evaluating the use of a CpG free promoter for long-term recombinant protein expression stability in Chinese hamster ovary cells.
Article in BMC biotechnology, 2016. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 10 papers.
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Who cites it
10 citing papers in PubMed, 22 citations in OpenAlex.
- From Cell Clones to Recombinant Protein Product Heterogeneity in Chinese Hamster Ovary Cell Systems.International journal of molecular sciences · 2025Review
- Evaluation of site-specific methylation of the CMV promoter and its role in CHO cell productivity of a recombinant monoclonal antibody.Antibody therapeutics · 2022Article
- Fluorescence-assisted sequential insertion of transgenes (FASIT): an approach for increasing specific productivity in mammalian cells.Scientific reports · 2020Article
- Expression vector cassette engineering for recombinant therapeutic production in mammalian cell systems.Applied microbiology and biotechnology · 2020Review
- Increased mAb production in amplified CHO cell lines is associated with increased interaction of CREB1 with transgene promoter.Current research in biotechnology · 2019Article
- Contribution of promoter DNA sequence to heterochromatin formation velocity and memory of gene repression in mouse embryo fibroblasts.PloS one · 2019Article
- CRISPR/Cas9-mediated gene knockout for DNA methyltransferase Dnmt3a in CHO cells displays enhanced transgenic expression and long-term stability.Journal of cellular and molecular medicine · 2018Article
- Blastocyst Formation Rate and Transgene Expression are Associated with Gene Insertion into Safe and Non-Safe Harbors in the Cattle Genome.Scientific reports · 2017Article
- Correlation Between Expression of Recombinant Proteins and Abundance of H3K4Me3 on the Enhancer of Human Cytomegalovirus Major Immediate-Early Promoter.Molecular biotechnology · 2017Article
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Authors and funding
6 authors at 3 institutions in 1 country.
Funding
No grant is acknowledged in the PubMed record.
Abstract
backgroundMethylated CpG dinucleotides in promoters are associated with the loss of gene expression in recombinant Chinese hamster ovary (CHO) cells during large-scale commercial manufacturing. We evaluated a promoter devoid of CpG dinucleotides, CpGfree, in parallel with a similar CpG containing promoter, CpGrich, for their ability to maintain the expression of recombinant enhanced green fluorescent protein (EGFP) after 8 weeks of culturing.
resultsWhile the promoters gave similar transient expression levels, CpGfree clones had significantly higher average stable expression possibly due to increased resistance to early silencing during integration into the chromosome. A greater proportion of cells in clones generated using the CpGfree promoter were still expressing detectable levels of EGFP after 8 weeks but the relative expression levels measured at week 8 to those measured at week 0 did not improve compared to clones generated using the CpGrich promoter. Chromatin immunoprecipitation assays indicated that the repression of the CpGfree promoter was likely linked to histone deacetylation and methylation. Use of histone deacetylase inhibitors also managed to recover some of the lost expression.
conclusionUsing a promoter without CpG dinucleotides could mitigate the early gene silencing but did not improve longer-term expression stability as silencing due to histone modifications could still take place. The results presented here would aid in promoter selection and design for improved protein production in CHO and other mammalian cells.
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