1. Cell Biology
  2. Neuroscience

A genetic toolkit for tagging intronic MiMIC containing genes

  1. Sonal Nagarkar-Jaiswal
  2. Steven Z DeLuca
  3. Pei-Tseng Lee
  4. Wen-Wen Lin
  5. Hongling Pan
  6. Zhongyuan Zuo
  7. Jiangxing Lv
  8. Allan C Spradling
  9. Hugo J Bellen  Is a corresponding author
  1. Howard Hughes Medical Institute, Baylor College of Medicine, United States
  2. Howard Hughes Medical Institute, Carnegie Institution for Science, United States
  3. Baylor College of Medicine, United States
https://doi.org/10.7554/eLife.08469
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Cite this article
  1. Sonal Nagarkar-Jaiswal
  2. Steven Z DeLuca
  3. Pei-Tseng Lee
  4. Wen-Wen Lin
  5. Hongling Pan
  6. Zhongyuan Zuo
  7. Jiangxing Lv
  8. Allan C Spradling
  9. Hugo J Bellen
(2015)
A genetic toolkit for tagging intronic MiMIC containing genes
eLife 4:e08469.
https://doi.org/10.7554/eLife.08469
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Abstract

Previously we described a large collection of MiMICs that contain two phiC31 recombinase target sites and allow the generation of a new exon that encodes a protein tag when the MiMIC (Minos Mediated Integration Cassette) is inserted in a codon intron (Nagarkar-Jaiswal et al., 2015). These modified genes permit numerous applications including assessment of protein expression pattern, identification of protein interaction partners by immunoprecipitation followed by mass spec, and reversible removal of the tagged protein in any tissue. At present, these conversions remain time and labor-intensive as they require embryos to be injected with plasmid DNA containing the exon tag. Here we describe a simple and reliable genetic strategy to tag genes/proteins that contain MiMIC insertions using an integrated exon encoding GFP flanked by FRT sequences. We document the efficiency and tag 60 mostly uncharacterized genes.

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Author details

  1. Sonal Nagarkar-Jaiswal

    Howard Hughes Medical Institute, Baylor College of Medicine, Houston, United States
    Competing interests
    The authors declare that no competing interests exist.

  2. Steven Z DeLuca

    Department of Embryology, Howard Hughes Medical Institute, Carnegie Institution for Science, Baltimore, United States
    Competing interests
    The authors declare that no competing interests exist.

  3. Pei-Tseng Lee

    Department of Molecular and Human Genetics, Baylor College of Medicine, Houston, United States
    Competing interests
    The authors declare that no competing interests exist.

  4. Wen-Wen Lin

    Department of Molecular and Human Genetics, Baylor College of Medicine, Houston, United States
    Competing interests
    The authors declare that no competing interests exist.

  5. Hongling Pan

    Howard Hughes Medical Institute, Baylor College of Medicine, Houston, United States
    Competing interests
    The authors declare that no competing interests exist.

  6. Zhongyuan Zuo

    Department of Molecular and Human Genetics, Baylor College of Medicine, Houston, United States
    Competing interests
    The authors declare that no competing interests exist.

  7. Jiangxing Lv

    Department of Molecular and Human Genetics, Baylor College of Medicine, Houston, United States
    Competing interests
    The authors declare that no competing interests exist.

  8. Allan C Spradling

    Department of Embryology, Howard Hughes Medical Institute, Carnegie Institution for Science, Baltimore, United States
    Competing interests
    The authors declare that no competing interests exist.

  9. Hugo J Bellen

    Howard Hughes Medical Institute, Baylor College of Medicine, Houston, United States
    For correspondence
    hbellen@bcm.edu
    Competing interests
    The authors declare that no competing interests exist.

Copyright

© 2015, Nagarkar-Jaiswal et al.

This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.

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  1. Sonal Nagarkar-Jaiswal
  2. Steven Z DeLuca
  3. Pei-Tseng Lee
  4. Wen-Wen Lin
  5. Hongling Pan
  6. Zhongyuan Zuo
  7. Jiangxing Lv
  8. Allan C Spradling
  9. Hugo J Bellen
(2015)
A genetic toolkit for tagging intronic MiMIC containing genes
eLife 4:e08469.
https://doi.org/10.7554/eLife.08469

Share this article

https://doi.org/10.7554/eLife.08469

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    1. Cell Biology
    2. Neuroscience

    A library of MiMICs allows tagging of genes and reversible, spatial and temporal knockdown of proteins in Drosophila

    Sonal Nagarkar-Jaiswal, Pei-Tseng Lee ... Hugo J Bellen
    Tools and Resources

    Here, we document a collection of ∼7434 MiMIC (Minos Mediated Integration Cassette) insertions of which 2854 are inserted in coding introns. They allowed us to create a library of 400 GFP-tagged genes. We show that 72% of internally tagged proteins are functional, and that more than 90% can be imaged in unfixed tissues. Moreover, the tagged mRNAs can be knocked down by RNAi against GFP (iGFPi), and the tagged proteins can be efficiently knocked down by deGradFP technology. The phenotypes associated with RNA and protein knockdown typically correspond to severe loss of function or null mutant phenotypes. Finally, we demonstrate reversible, spatial, and temporal knockdown of tagged proteins in larvae and adult flies. This new strategy and collection of strains allows unprecedented in vivo manipulations in flies for many genes. These strategies will likely extend to vertebrates.