A Conditional System to Specifically Link Disruption of Protein-Coding Function with Reporter Expression in Mice
Conditional gene deletion in mice has contributed immensely to our understanding of many biological and biomedical processes. Despite an increasing awareness of nonprotein-coding functional elements within protein-coding transcripts, current gene-targeting approaches typically involve simultaneous a...
| Main Authors: | , , , , , , , , , |
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| Format: | Article |
| Language: | en_US |
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Elsevier B.V.
2015
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| Online Access: | http://hdl.handle.net/1721.1/96530 https://orcid.org/0000-0001-5785-8911 |
| _version_ | 1826201611432624128 |
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| author | Chiou, Shin-Heng Kim-Kiselak, Caroline Risca, Viviana I. Heimann, Megan Chuang, Chen-Hua Burds, Aurora A. Greenleaf, William J. Jacks, Tyler E. Feldser, David M. Winslow, Monte M. |
| author2 | Koch Institute for Integrative Cancer Research at MIT |
| author_facet | Koch Institute for Integrative Cancer Research at MIT Chiou, Shin-Heng Kim-Kiselak, Caroline Risca, Viviana I. Heimann, Megan Chuang, Chen-Hua Burds, Aurora A. Greenleaf, William J. Jacks, Tyler E. Feldser, David M. Winslow, Monte M. |
| author_sort | Chiou, Shin-Heng |
| collection | MIT |
| description | Conditional gene deletion in mice has contributed immensely to our understanding of many biological and biomedical processes. Despite an increasing awareness of nonprotein-coding functional elements within protein-coding transcripts, current gene-targeting approaches typically involve simultaneous ablation of noncoding elements within targeted protein-coding genes. The potential for protein-coding genes to have additional noncoding functions necessitates the development of novel genetic tools capable of precisely interrogating individual functional elements. We present a strategy that couples Cre/loxP-mediated conditional gene disruption with faithful GFP reporter expression in mice in which Cre-mediated stable inversion of a splice acceptor-GFP-splice donor cassette concurrently disrupts protein production and creates a GFP fusion product. Importantly, cassette inversion maintains physiologic transcript structure, thereby ensuring proper microRNA-mediated regulation of the GFP reporter, as well as maintaining expression of nonprotein-coding elements. To test this potentially generalizable strategy, we generated and analyzed mice with this conditional knockin reporter targeted to the Hmga2 locus. |
| first_indexed | 2024-09-23T11:54:23Z |
| format | Article |
| id | mit-1721.1/96530 |
| institution | Massachusetts Institute of Technology |
| language | en_US |
| last_indexed | 2024-09-23T11:54:23Z |
| publishDate | 2015 |
| publisher | Elsevier B.V. |
| record_format | dspace |
| spelling | mit-1721.1/965302022-10-01T06:50:18Z A Conditional System to Specifically Link Disruption of Protein-Coding Function with Reporter Expression in Mice Chiou, Shin-Heng Kim-Kiselak, Caroline Risca, Viviana I. Heimann, Megan Chuang, Chen-Hua Burds, Aurora A. Greenleaf, William J. Jacks, Tyler E. Feldser, David M. Winslow, Monte M. Koch Institute for Integrative Cancer Research at MIT Kim-Kiselak, Caroline Heimann, Megan Burds, Aurora A. Jacks, Tyler E. Conditional gene deletion in mice has contributed immensely to our understanding of many biological and biomedical processes. Despite an increasing awareness of nonprotein-coding functional elements within protein-coding transcripts, current gene-targeting approaches typically involve simultaneous ablation of noncoding elements within targeted protein-coding genes. The potential for protein-coding genes to have additional noncoding functions necessitates the development of novel genetic tools capable of precisely interrogating individual functional elements. We present a strategy that couples Cre/loxP-mediated conditional gene disruption with faithful GFP reporter expression in mice in which Cre-mediated stable inversion of a splice acceptor-GFP-splice donor cassette concurrently disrupts protein production and creates a GFP fusion product. Importantly, cassette inversion maintains physiologic transcript structure, thereby ensuring proper microRNA-mediated regulation of the GFP reporter, as well as maintaining expression of nonprotein-coding elements. To test this potentially generalizable strategy, we generated and analyzed mice with this conditional knockin reporter targeted to the Hmga2 locus. Stanford University. School of Medicine (Dean’s postdoctoral fellow) American Lung Association (Fellowship) National Institutes of Health (U.S.) (Stanford Cancer Institute support grant (P30- CA124435)) National Institutes of Health (U.S.) (MIT Cancer Center support grant (P30-CA14051)) National Institutes of Health (U.S.) (grant R00-CA151968) David H. Koch Institute for Integrative Cancer Research at MIT (Daniel K. Ludwig Scholar) Howard Hughes Medical Institute (Investigator) American Association for Cancer Research (Pancreatic Cancer Action Network-AACR Career Development Awards, in memory of Skip Viragh (13- 20-25-WINS)) National Institutes of Health (U.S.) (grant R00-CA158581) Stanford University (Walter V. and Idun Berry Postdoctoral Fellowship) 2015-04-13T15:24:58Z 2015-04-13T15:24:58Z 2014-06 2014-04 Article http://purl.org/eprint/type/JournalArticle 22111247 http://hdl.handle.net/1721.1/96530 Chiou, Shin-Heng, Caroline Kim-Kiselak, Viviana I. Risca, Megan K. Heimann, Chen-Hua Chuang, Aurora A. Burds, William J. Greenleaf, Tyler E. Jacks, David M. Feldser, and Monte M. Winslow. “A Conditional System to Specifically Link Disruption of Protein-Coding Function with Reporter Expression in Mice.” Cell Reports 7, no. 6 (June 2014): 2078–2086. https://orcid.org/0000-0001-5785-8911 en_US http://dx.doi.org/10.1016/j.celrep.2014.05.031 Cell Reports Creative Commons Attribution-NonCommercial-NoDerivs 3.0 License http://creativecommons.org/licenses/by-nc-nd/3.0/ application/pdf Elsevier B.V. Elsevier Open Access |
| spellingShingle | Chiou, Shin-Heng Kim-Kiselak, Caroline Risca, Viviana I. Heimann, Megan Chuang, Chen-Hua Burds, Aurora A. Greenleaf, William J. Jacks, Tyler E. Feldser, David M. Winslow, Monte M. A Conditional System to Specifically Link Disruption of Protein-Coding Function with Reporter Expression in Mice |
| title | A Conditional System to Specifically Link Disruption of Protein-Coding Function with Reporter Expression in Mice |
| title_full | A Conditional System to Specifically Link Disruption of Protein-Coding Function with Reporter Expression in Mice |
| title_fullStr | A Conditional System to Specifically Link Disruption of Protein-Coding Function with Reporter Expression in Mice |
| title_full_unstemmed | A Conditional System to Specifically Link Disruption of Protein-Coding Function with Reporter Expression in Mice |
| title_short | A Conditional System to Specifically Link Disruption of Protein-Coding Function with Reporter Expression in Mice |
| title_sort | conditional system to specifically link disruption of protein coding function with reporter expression in mice |
| url | http://hdl.handle.net/1721.1/96530 https://orcid.org/0000-0001-5785-8911 |
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