ArticleDisease models & mechanisms2011
Quantification of factors influencing fluorescent protein expression using RMCE to generate an allelic series in the ROSA26 locus in mice.
Article in Disease models & mechanisms, 2011. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 38 papers.
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Who cites it
38 citing papers in PubMed, 51 citations in OpenAlex.
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- Transgene-associated human growth hormone expression in pancreatic β-cells impairs identification of sex-based gene expression differences.American journal of physiology. Endocrinology and metabolism · 2019Article
- Pancreatic islet-autonomous insulin and smoothened-mediated signalling modulate identity changes of glucagonNature cell biology · 2018Article
- Site-specific chromosomal gene insertion: Flp recombinase versus Cas9 nuclease.Scientific reports · 2017Article
- Efficiency and Specificity of Gene Deletion in Lung Epithelial Doxycycline-Inducible Cre Mice.American journal of respiratory cell and molecular biology · 2017Article
- Imaging morphogenesis.Philosophical transactions of the Royal Society of London. Series B, Biological sciences · 2017Review
- Optical Barcoding for Single-Clone Tracking to Study Tumor Heterogeneity.Molecular therapy : the journal of the American Society of Gene Therapy · 2017Article
- Setd5 is essential for mammalian development and the co-transcriptional regulation of histone acetylation.Development (Cambridge, England) · 2016Article
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Authors and funding
9 authors at 1 institution in 1 country.
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Abstract
Fluorescent proteins (FPs) have great utility in identifying specific cell populations and in studying cellular dynamics in the mouse. To quantify the factors that determine both the expression and relative brightness of FPs in mouse embryonic stem cells (mESCs) and in mice, we generated eight different FP-expressing ROSA26 alleles using recombinase-mediated cassette exchange (RMCE). These alleles enabled us to analyze the effects on FP expression of a translational enhancer and different 3'-intronic and/or polyadenylation sequences, as well as the relative brightness of five different FPs, without the confounding position and copy number effects that are typically associated with randomly inserted transgenes. We found that the expression of a given FP can vary threefold or more depending on the genetic features present in the allele. The optimal FP expression cassette contained both a translational enhancer sequence in the 5'-untranslated region (UTR) and an intron-containing rabbit β-globin sequence within the 3'-UTR. The relative expressed brightness of individual FPs varied up to tenfold. Of the five different monomeric FPs tested, Citrine (YFP) was the brightest, followed by Apple, eGFP, Cerulean (CFP) and Cherry. Generation of a line of Cherry-expressing mice showed that there was a 30-fold variation of Cherry expression among different tissues and that there was a punctate expression pattern within cells of all tissues examined. This study should help investigators make better-informed design choices when expressing FPs in mESCs and mice.
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