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A ratiometric dual luciferase reporter for quantitative monitoring of pre-mRNA splicing efficiency in vivo
Precursor messenger RNA (pre-mRNA) splicing is critical for cell growth and development, and errors in RNA splicing frequently cause cellular dysfunction, abnormal gene expression, and a variety of human diseases. However, there is currently a lack of reliable systems to noninvasively monitor the mR...
Autores principales: | , , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
American Society for Biochemistry and Molecular Biology
2021
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8322121/ https://www.ncbi.nlm.nih.gov/pubmed/34216622 http://dx.doi.org/10.1016/j.jbc.2021.100933 |
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author | Guo, Bin Shi, Xiaorui Ma, Zhe Ji, Moxuan Tang, Chu Wang, Fu |
author_facet | Guo, Bin Shi, Xiaorui Ma, Zhe Ji, Moxuan Tang, Chu Wang, Fu |
author_sort | Guo, Bin |
collection | PubMed |
description | Precursor messenger RNA (pre-mRNA) splicing is critical for cell growth and development, and errors in RNA splicing frequently cause cellular dysfunction, abnormal gene expression, and a variety of human diseases. However, there is currently a lack of reliable systems to noninvasively monitor the mRNA splicing efficiency in cells and animals. Here, we described the design of a genetically engineered ratiometric dual luciferase reporter to continuously quantify the changes in mRNA splice variants in vivo. This reporter system is encoded within a single polypeptide but on separate exons, thus generating two distinct luciferase signals derived from spliced and unspliced mRNAs. With this reporter, the two kinds of luciferase in the same individual can minimize the influence of indirect factors on splicing, and the ratio of these two luciferase intensities represents the dynamic splicing efficiency of pre-mRNA. Our study offers a convenient and robust tool for the screening and identification of small molecules or trans-acting factors that affect the efficiency of specific splicing reactions. |
format | Online Article Text |
id | pubmed-8322121 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2021 |
publisher | American Society for Biochemistry and Molecular Biology |
record_format | MEDLINE/PubMed |
spelling | pubmed-83221212021-07-31 A ratiometric dual luciferase reporter for quantitative monitoring of pre-mRNA splicing efficiency in vivo Guo, Bin Shi, Xiaorui Ma, Zhe Ji, Moxuan Tang, Chu Wang, Fu J Biol Chem Accelerated Communication Precursor messenger RNA (pre-mRNA) splicing is critical for cell growth and development, and errors in RNA splicing frequently cause cellular dysfunction, abnormal gene expression, and a variety of human diseases. However, there is currently a lack of reliable systems to noninvasively monitor the mRNA splicing efficiency in cells and animals. Here, we described the design of a genetically engineered ratiometric dual luciferase reporter to continuously quantify the changes in mRNA splice variants in vivo. This reporter system is encoded within a single polypeptide but on separate exons, thus generating two distinct luciferase signals derived from spliced and unspliced mRNAs. With this reporter, the two kinds of luciferase in the same individual can minimize the influence of indirect factors on splicing, and the ratio of these two luciferase intensities represents the dynamic splicing efficiency of pre-mRNA. Our study offers a convenient and robust tool for the screening and identification of small molecules or trans-acting factors that affect the efficiency of specific splicing reactions. American Society for Biochemistry and Molecular Biology 2021-07-01 /pmc/articles/PMC8322121/ /pubmed/34216622 http://dx.doi.org/10.1016/j.jbc.2021.100933 Text en © 2021 The Authors https://creativecommons.org/licenses/by-nc-nd/4.0/This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/4.0/). |
spellingShingle | Accelerated Communication Guo, Bin Shi, Xiaorui Ma, Zhe Ji, Moxuan Tang, Chu Wang, Fu A ratiometric dual luciferase reporter for quantitative monitoring of pre-mRNA splicing efficiency in vivo |
title | A ratiometric dual luciferase reporter for quantitative monitoring of pre-mRNA splicing efficiency in vivo |
title_full | A ratiometric dual luciferase reporter for quantitative monitoring of pre-mRNA splicing efficiency in vivo |
title_fullStr | A ratiometric dual luciferase reporter for quantitative monitoring of pre-mRNA splicing efficiency in vivo |
title_full_unstemmed | A ratiometric dual luciferase reporter for quantitative monitoring of pre-mRNA splicing efficiency in vivo |
title_short | A ratiometric dual luciferase reporter for quantitative monitoring of pre-mRNA splicing efficiency in vivo |
title_sort | ratiometric dual luciferase reporter for quantitative monitoring of pre-mrna splicing efficiency in vivo |
topic | Accelerated Communication |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8322121/ https://www.ncbi.nlm.nih.gov/pubmed/34216622 http://dx.doi.org/10.1016/j.jbc.2021.100933 |
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