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Massively Parallel Reporter Assays for High-Throughput In Vivo Analysis of Cis-Regulatory Elements

The rapid improvement of descriptive genomic technologies has fueled a dramatic increase in hypothesized connections between cardiovascular gene expression and phenotypes. However, in vivo testing of these hypotheses has predominantly been relegated to slow, expensive, and linear generation of genet...

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Autores principales: Zheng, Yanjiang, VanDusen, Nathan J.
Formato: Online Artículo Texto
Lenguaje:English
Publicado: MDPI 2023
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10146671/
https://www.ncbi.nlm.nih.gov/pubmed/37103023
http://dx.doi.org/10.3390/jcdd10040144
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author Zheng, Yanjiang
VanDusen, Nathan J.
author_facet Zheng, Yanjiang
VanDusen, Nathan J.
author_sort Zheng, Yanjiang
collection PubMed
description The rapid improvement of descriptive genomic technologies has fueled a dramatic increase in hypothesized connections between cardiovascular gene expression and phenotypes. However, in vivo testing of these hypotheses has predominantly been relegated to slow, expensive, and linear generation of genetically modified mice. In the study of genomic cis-regulatory elements, generation of mice featuring transgenic reporters or cis-regulatory element knockout remains the standard approach. While the data obtained is of high quality, the approach is insufficient to keep pace with candidate identification and therefore results in biases introduced during the selection of candidates for validation. However, recent advances across a range of disciplines are converging to enable functional genomic assays that can be conducted in a high-throughput manner. Here, we review one such method, massively parallel reporter assays (MPRAs), in which the activities of thousands of candidate genomic regulatory elements are simultaneously assessed via the next-generation sequencing of a barcoded reporter transcript. We discuss best practices for MPRA design and use, with a focus on practical considerations, and review how this emerging technology has been successfully deployed in vivo. Finally, we discuss how MPRAs are likely to evolve and be used in future cardiovascular research.
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spelling pubmed-101466712023-04-29 Massively Parallel Reporter Assays for High-Throughput In Vivo Analysis of Cis-Regulatory Elements Zheng, Yanjiang VanDusen, Nathan J. J Cardiovasc Dev Dis Review The rapid improvement of descriptive genomic technologies has fueled a dramatic increase in hypothesized connections between cardiovascular gene expression and phenotypes. However, in vivo testing of these hypotheses has predominantly been relegated to slow, expensive, and linear generation of genetically modified mice. In the study of genomic cis-regulatory elements, generation of mice featuring transgenic reporters or cis-regulatory element knockout remains the standard approach. While the data obtained is of high quality, the approach is insufficient to keep pace with candidate identification and therefore results in biases introduced during the selection of candidates for validation. However, recent advances across a range of disciplines are converging to enable functional genomic assays that can be conducted in a high-throughput manner. Here, we review one such method, massively parallel reporter assays (MPRAs), in which the activities of thousands of candidate genomic regulatory elements are simultaneously assessed via the next-generation sequencing of a barcoded reporter transcript. We discuss best practices for MPRA design and use, with a focus on practical considerations, and review how this emerging technology has been successfully deployed in vivo. Finally, we discuss how MPRAs are likely to evolve and be used in future cardiovascular research. MDPI 2023-03-29 /pmc/articles/PMC10146671/ /pubmed/37103023 http://dx.doi.org/10.3390/jcdd10040144 Text en © 2023 by the authors. https://creativecommons.org/licenses/by/4.0/Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (https://creativecommons.org/licenses/by/4.0/).
spellingShingle Review
Zheng, Yanjiang
VanDusen, Nathan J.
Massively Parallel Reporter Assays for High-Throughput In Vivo Analysis of Cis-Regulatory Elements
title Massively Parallel Reporter Assays for High-Throughput In Vivo Analysis of Cis-Regulatory Elements
title_full Massively Parallel Reporter Assays for High-Throughput In Vivo Analysis of Cis-Regulatory Elements
title_fullStr Massively Parallel Reporter Assays for High-Throughput In Vivo Analysis of Cis-Regulatory Elements
title_full_unstemmed Massively Parallel Reporter Assays for High-Throughput In Vivo Analysis of Cis-Regulatory Elements
title_short Massively Parallel Reporter Assays for High-Throughput In Vivo Analysis of Cis-Regulatory Elements
title_sort massively parallel reporter assays for high-throughput in vivo analysis of cis-regulatory elements
topic Review
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10146671/
https://www.ncbi.nlm.nih.gov/pubmed/37103023
http://dx.doi.org/10.3390/jcdd10040144
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