Design of a GFP reporter for splicing analysis in mammalian cells
Eukaryotic genes are formed by exons and introns. Pre-mRNA splicing promotes exon ligation and intron removal and is performed by a specialized macromolecular machinery named spliceosome, composed of five small ribonucleoprotein particles (snRNPs) and more than one hundred proteins. The activity of...
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| Language: | English |
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Elsevier
2025-06-01
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| Series: | Biotechnology Reports |
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| Online Access: | http://www.sciencedirect.com/science/article/pii/S2215017X25000141 |
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| author | Arthur T. Menezes Helder Y. Nagasse Hilan R. M. Lopes Patricia P. Coltri |
| author_facet | Arthur T. Menezes Helder Y. Nagasse Hilan R. M. Lopes Patricia P. Coltri |
| author_sort | Arthur T. Menezes |
| collection | DOAJ |
| description | Eukaryotic genes are formed by exons and introns. Pre-mRNA splicing promotes exon ligation and intron removal and is performed by a specialized macromolecular machinery named spliceosome, composed of five small ribonucleoprotein particles (snRNPs) and more than one hundred proteins. The activity of this complex is highly accurate due to the coordinated activity of its components. Altered splicing has been related to the development of several diseases, including neurodegenerative disorders, such as amyotrophic lateral sclerosis, and different types of cancer. Detailed understanding of splicing regulation in eukaryotic cells can be achieved using splicing reporter systems. We designed a reporter plasmid suitable for splicing analysis in cultured mammalian cells. Our reporter is based on GFP expression, and the splicing outcome can be easily visualized by fluorescence microscopy. We quantified splicing activity in two human cell lines, HEK-293T and MDA-MB-231, confirming its suitability for use in live cells in culture. |
| format | Article |
| id | doaj-art-6c6e6ba0f0d941bfa18bee94f5256243 |
| institution | DOAJ |
| issn | 2215-017X |
| language | English |
| publishDate | 2025-06-01 |
| publisher | Elsevier |
| record_format | Article |
| series | Biotechnology Reports |
| spelling | doaj-art-6c6e6ba0f0d941bfa18bee94f52562432025-08-20T03:05:54ZengElsevierBiotechnology Reports2215-017X2025-06-0146e0088710.1016/j.btre.2025.e00887Design of a GFP reporter for splicing analysis in mammalian cellsArthur T. Menezes0Helder Y. Nagasse1Hilan R. M. Lopes2Patricia P. Coltri3Department of Cell and Developmental Biology, Institute of Biomedical Sciences, University of São Paulo, 05508-000, BrazilDepartment of Cell and Developmental Biology, Institute of Biomedical Sciences, University of São Paulo, 05508-000, BrazilDepartment of Cell and Developmental Biology, Institute of Biomedical Sciences, University of São Paulo, 05508-000, BrazilCorresponding author.; Department of Cell and Developmental Biology, Institute of Biomedical Sciences, University of São Paulo, 05508-000, BrazilEukaryotic genes are formed by exons and introns. Pre-mRNA splicing promotes exon ligation and intron removal and is performed by a specialized macromolecular machinery named spliceosome, composed of five small ribonucleoprotein particles (snRNPs) and more than one hundred proteins. The activity of this complex is highly accurate due to the coordinated activity of its components. Altered splicing has been related to the development of several diseases, including neurodegenerative disorders, such as amyotrophic lateral sclerosis, and different types of cancer. Detailed understanding of splicing regulation in eukaryotic cells can be achieved using splicing reporter systems. We designed a reporter plasmid suitable for splicing analysis in cultured mammalian cells. Our reporter is based on GFP expression, and the splicing outcome can be easily visualized by fluorescence microscopy. We quantified splicing activity in two human cell lines, HEK-293T and MDA-MB-231, confirming its suitability for use in live cells in culture.http://www.sciencedirect.com/science/article/pii/S2215017X25000141Pre-mRNA splicingGFPReporter system |
| spellingShingle | Arthur T. Menezes Helder Y. Nagasse Hilan R. M. Lopes Patricia P. Coltri Design of a GFP reporter for splicing analysis in mammalian cells Biotechnology Reports Pre-mRNA splicing GFP Reporter system |
| title | Design of a GFP reporter for splicing analysis in mammalian cells |
| title_full | Design of a GFP reporter for splicing analysis in mammalian cells |
| title_fullStr | Design of a GFP reporter for splicing analysis in mammalian cells |
| title_full_unstemmed | Design of a GFP reporter for splicing analysis in mammalian cells |
| title_short | Design of a GFP reporter for splicing analysis in mammalian cells |
| title_sort | design of a gfp reporter for splicing analysis in mammalian cells |
| topic | Pre-mRNA splicing GFP Reporter system |
| url | http://www.sciencedirect.com/science/article/pii/S2215017X25000141 |
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