LINE-1 Cultured Cell Retrotransposition Assay

Huira C Kopera; Peter A Larson; John B Moldovan; Sandra R Richardson; Ying Liu; John V Moran

doi:10.1007/978-1-4939-3372-3_10

LINE-1 Cultured Cell Retrotransposition Assay

Methods Mol Biol. 2016:1400:139-56. doi: 10.1007/978-1-4939-3372-3_10.

Authors

Huira C Kopera¹, Peter A Larson², John B Moldovan³, Sandra R Richardson², Ying Liu⁴, John V Moran^{5

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Affiliations

¹ Department of Human Genetics, University of Michigan Medical School, 1241 E. Catherine St, Ann Arbor, MI, 48109, USA. chongh@umich.edu.
² Department of Human Genetics, University of Michigan Medical School, 1241 E. Catherine St, Ann Arbor, MI, 48109, USA.
³ Cellular and Molecular Biology Program, University of Michigan Medical School, 1241 E. Catherine St, Ann Arbor, MI, 48109, USA.
⁴ Howard Hughes Medical Institute, Chevy Chase, MD, USA.
⁵ Department of Human Genetics, University of Michigan Medical School, 1241 E. Catherine St, Ann Arbor, MI, 48109, USA. moranj@umich.edu.
⁶ Cellular and Molecular Biology Program, University of Michigan Medical School, Ann Arbor, MI, 48109, USA. moranj@umich.edu.
⁷ Department of Internal Medicine, University of Michigan Medical School, Ann Arbor, MI, 48109, USA. moranj@umich.edu.
⁸ Howard Hughes Medical Institute, Chevy Chase, MD, USA. moranj@umich.edu.

Abstract

The Long INterspersed Element-1 (LINE-1 or L1) retrotransposition assay has facilitated the discovery and characterization of active (i.e., retrotransposition-competent) LINE-1 sequences from mammalian genomes. In this assay, an engineered LINE-1 containing a retrotransposition reporter cassette is transiently transfected into a cultured cell line. Expression of the reporter cassette, which occurs only after a successful round of retrotransposition, allows the detection and quantification of the LINE-1 retrotransposition efficiency. This assay has yielded insight into the mechanism of LINE-1 retrotransposition. It also has provided a greater understanding of how the cell regulates LINE-1 retrotransposition and how LINE-1 retrotransposition impacts the structure of mammalian genomes. Below, we provide a brief introduction to LINE-1 biology and then detail how the LINE-1 retrotransposition assay is performed in cultured mammalian cells.

Keywords: Alu; LINE-1; Mammalian cultured cells; Retrotransposition assay; trans-complementation assay.

Publication types

Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't

MeSH terms

Alu Elements
Gene Expression
HeLa Cells
Humans
Long Interspersed Nucleotide Elements*
Retroelements
Transfection

Substances

Retroelements

Abstract

Publication types

MeSH terms

Substances

Grants and funding