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Cited by 98Chromosomal and Genetic VariationsGenomic variations and chromosomal abnormalitiesGenome Rearrangement Algorithms

Characterization of L1 retrotransposition with high-throughput dual-luciferase assays

Open access
Xie, Yi, Rosser, James M., Thompson, Tina L., Boeke, Jef D., An, Wenfeng·2011·Nucleic Acids Research
DOI 10.1093/nar/gkq1076PMC PMC3035435
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Abstract

Recent studies employing genome-wide approaches have provided an unprecedented view of the scope of L1 activities on structural variations in the human genome, and further reinforced the role of L1s as one of the major driving forces behind human genome evolution. The rapid identification of novel L1 elements by these high-throughput approaches demands improved L1 functional assays. However, the existing assays use antibiotic selection markers or fluorescent proteins as reporters; neither is amenable to miniaturization. To increase assay sensitivity and throughput, we have developed a third generation assay by using dual-luciferase reporters, in which firefly luciferase is used as the retrotransposition indicator and Renilla luciferase is encoded on the same or separate plasmid for normalization. This novel assay is highly sensitive and has a broad dynamic range. Quantitative data with high signal-to-noise ratios can be obtained from 24- up to 96-well plates in 2-4 days after transfection. Using the dual-luciferase assays, we have characterized profiles of retrotransposition by various human and mouse L1 elements, and detailed the kinetics of L1 retrotransposition in cultured cells. Its high-throughput and short assay timeframe make it well suited for routine tests as well as large-scale screening efforts.

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Crossref · 10.1093/nar/gkq1076

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  • pWA366 contains L1RP(JM111), a mutated element carrying two missense mutations in ORF1 that is unable to retrotranspose.

    “pWA355 contains a functional, active L1RP element, whereas pWA366 contains L1RP(JM111), a mutated element carrying two missense mutations in ORF1 that is unable to retrotranspose.”

    From L1 drives IFN in senescent cells and promotes age-associated inflammation (2019) · cites this paper as Xie et al. (2011)52

  • pWA355 contains a functional, active L1RP element.

    “pWA355 contains a functional, active L1RP element, whereas pWA366 contains L1RP(JM111), a mutated element carrying two missense mutations in ORF1 that is unable to retrotranspose.”

    From L1 drives IFN in senescent cells and promotes age-associated inflammation (2019) · cites this paper as Xie et al. (2011)52

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Cites

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Research works printed but not linked: 26

  1. Lander, 2001, Initial sequencing and analysis of the human genome, Nature, 409, 860, 10.1038/35057062 · DOI · Paper page
  2. Kazazian, 2004, Mobile elements: drivers of genome evolution, Science, 303, 1626, 10.1126/science.1089670 · DOI · Paper page
  3. Cordaux, 2009, The impact of retrotransposons on human genome evolution, Nat. Rev. Genet., 10, 691, 10.1038/nrg2640 · DOI
  4. Esnault, 2000, Human LINE retrotransposons generate processed pseudogenes, Nat. Genet., 24, 363, 10.1038/74184 · DOI
  5. Wei, 2001, Human L1 retrotransposition: cis preference versus trans complementation, Mol. Cell Biol., 21, 1429, 10.1128/MCB.21.4.1429-1439.2001 · DOI
  6. Dewannieux, 2003, LINE-mediated retrotransposition of marked Alu sequences, Nat. Genet., 35, 41, 10.1038/ng1223 · DOI
  7. Ostertag, 2003, SVA elements are nonautonomous retrotransposons that cause disease in humans, Am. J. Hum. Genet., 73, 1444, 10.1086/380207 · DOI
  8. Wang, 2005, SVA elements: a hominid-specific retroposon family, J. Mol. Biol., 354, 994, 10.1016/j.jmb.2005.09.085 · DOI
  9. Belancio, 2008, Mammalian non-LTR retrotransposons: for better or worse, in sickness and in health, Genome Res., 18, 343, 10.1101/gr.5558208 · DOI
  10. Chen, 2005, A systematic analysis of LINE-1 endonuclease-dependent retrotranspositional events causing human genetic disease, Hum. Genet., 117, 411, 10.1007/s00439-005-1321-0 · DOI
  11. Brouha, 2003, Hot L1s account for the bulk of retrotransposition in the human population, Proc. Natl Acad. Sci. USA, 100, 5280, 10.1073/pnas.0831042100 · DOI · Paper page
  12. Huang, 2010, Mobile interspersed repeats are major structural variants in the human genome, Cell, 141, 1171, 10.1016/j.cell.2010.05.026 · DOI
  13. Ewing, 2010, High-throughput sequencing reveals extensive variation in human-specific L1 content in individual human genomes, Genome Res., 20, 1262, 10.1101/gr.106419.110 · DOI
  14. Iskow, 2010, Natural mutagenesis of human genomes by endogenous retrotransposons, Cell, 141, 1253, 10.1016/j.cell.2010.05.020 · DOI · Paper page
  15. Moran, 1996, High frequency retrotransposition in cultured mammalian cells, Cell, 87, 917, 10.1016/S0092-8674(00)81998-4 · DOI · Paper page
  16. Rangwala, 2009, The L1 retrotransposition assay: a retrospective and toolkit, Methods, 49, 219, 10.1016/j.ymeth.2009.04.012 · DOI
  17. Wei, 2000, A transient assay reveals that cultured human cells can accommodate multiple LINE-1 retrotransposition events, Anal. Biochem., 284, 435, 10.1006/abio.2000.4675 · DOI
  18. An, 2009, Plug and play modular strategies for synthetic retrotransposons, Methods, 49, 227, 10.1016/j.ymeth.2009.05.013 · DOI
  19. Rosser, 2010, Repeat-induced gene silencing of L1 transgenes is correlated with differential promoter methylation, Gene, 456, 15, 10.1016/j.gene.2010.02.005 · DOI
  20. Kimberland, 1999, Full-length human L1 insertions retain the capacity for high frequency retrotransposition in cultured cells, Hum. Mol. Genet., 8, 1557, 10.1093/hmg/8.8.1557 · DOI
  21. An, 2006, Active retrotransposition by a synthetic L1 element in mice, Proc. Natl Acad. Sci. USA, 103, 18662, 10.1073/pnas.0605300103 · DOI
  22. Han, 2004, A highly active synthetic mammalian retrotransposon, Nature, 429, 314, 10.1038/nature02535 · DOI
  23. Grimaldi, 1984, Defining the beginning and end of KpnI family segments, EMBO J., 3, 1753, 10.1002/j.1460-2075.1984.tb02042.x · DOI
  24. Kroutter, 2009, The RNA polymerase dictates ORF1 requirement and timing of LINE and SINE retrotransposition, PLoS Genet., 5, e1000458, 10.1371/journal.pgen.1000458 · DOI
  25. Garcia-Perez, 2010, Epigenetic silencing of engineered L1 retrotransposition events in human embryonic carcinoma cells, Nature, 466, 769, 10.1038/nature09209 · DOI
  26. Belancio, 2006, LINE-1 RNA splicing and influences on mammalian gene expression, Nucleic Acids Res., 34, 1512, 10.1093/nar/gkl027 · DOI

Cited by

Sorted by citations

Authors
Xie, Y., Rosser, J. M., Thompson, T. L., Boeke, J. D., & An, W.
Affiliations
Johns Hopkins University (US)
Corresponding author
An, Wenfeng
ORCID
Venue
Nucleic Acids Research
Publisher
Oxford University Press (OUP)
Published
2010-11-10
Volume
39
Issue
3
Pages
e16-e16
ISSN
  • 0305-1048 (ISSN-L)
  • 1362-4962
DOI
10.1093/nar/gkq1076
PMC
PMC3035435
OpenAlex
W2115594110
Cited by
98
References
3
Citations by year
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FWCI
2.69
Type
journal article
Language
en
Open access
Licence
Locations
  • Nucleic Acids Research (publishedVersion, cc-by-nc)
  • PubMed Central (publishedVersion, cc-by-nc)
  • PubMed (publishedVersion)
Funders
  • Washington State University
  • National Cancer Institute (P01CA16519, P01CA016519)
Topics
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Genetics › Chromosomal and Genetic Variations
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Genetics › Genomic variations and chromosomal abnormalities
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Genetics › Genome Rearrangement Algorithms
Keywords
L1 retrotransposition, long interspersed nuclear elements, dual-luciferase reporter assay, firefly luciferase, Renilla luciferase, structural variants, human genome evolution, high-throughput functional assay, miniaturization
MeSH terms
Animals, Genetic Vectors, HeLa Cells, Humans, Introns, Kinetics, Miniaturization, Promoter Regions, Genetic, Genes, Reporter, Reverse Transcriptase Inhibitors, Long Interspersed Nucleotide Elements, Luminescent Agents, Luciferases, Firefly, Luciferases, Renilla, Mice

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