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A Method for Inducible Gene Over-Expression and Down-Regulation in Emerging Model Species Using Pogostick

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Gene Function Analysis

Part of the book series: Methods in Molecular Biology ((MIMB,volume 1101))

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Abstract

Nontraditional model species need new tools for the functional testing of genes, both conserved and lineage-specific genes. These tools should enable the exploration of gene function, either via knock-downs of endogenous genes or via over-expression and ectopic expression of transgenes. We constructed a new vector called Pogostick that can be used to over-express or down-regulate genes in organisms amenable to germ-line transformation by the piggyBac transposable element. The vector currently uses the heat-shock promoter Hsp70 from Drosophila melanogaster to drive transgene expression and, as such, will have immediate applicability to organisms that can correctly interpret this promotor sequence. Here we introduce the main features of Pogostick and how candidate genes can be inserted into the vector for use in either over-expression or down-regulation experiments. In addition, we also test Pogostick in two insect species, D. melanogaster and the emerging model butterfly Bicyclus anynana. We over-express the fluorescent protein DsRed during the larval and pupal stages of D. melanogaster development, and down-regulate DsRed in a line constitutively expressing this gene in the eyes. We then test the over-expression of Ultrabithorax (Ubx) in B. anynana, and obtain sequences flanking the Pogostick genomic insertions. This new vector will allow emerging model species to enter the field of functional genetics with few hurdles.

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Acknowledgments

This work was supported by grants from The National Natural Science Foundation of China (31071968 and 31372265), grants from the National Institute of Health (R01 AI095184), Key Scientific and Technological Project of Chongqing (CSTC2012GG-YYJSB80002) and Par-Eu Scholars Program.

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Authors and Affiliations

  1. Institute of Entomology and Molecular Biology, College of Life Sciences, Chongqing Normal University, Chongqing, P. R. China

    Bin Chen

  2. Department of Ecology and Evolutionary Biology, Yale University, New Haven, CT, USA

    Antónia Monteiro

Authors
  1. Bin Chen
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  2. Antónia Monteiro
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Editors and Affiliations

  1. Dept of Mathematics & Statistics, The College of New Jersey, Ewing, New Jersey, USA

    Michael F. Ochs

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Chen, B., Monteiro, A. (2014). A Method for Inducible Gene Over-Expression and Down-Regulation in Emerging Model Species Using Pogostick . In: Ochs, M. (eds) Gene Function Analysis. Methods in Molecular Biology, vol 1101. Humana Press, Totowa, NJ. https://doi.org/10.1007/978-1-62703-721-1_12

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  • DOI: https://doi.org/10.1007/978-1-62703-721-1_12

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  • Publisher Name: Humana Press, Totowa, NJ

  • Print ISBN: 978-1-62703-720-4

  • Online ISBN: 978-1-62703-721-1

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