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Engineering rules that minimize germline silencing of transgenes in simple extrachromosomal arrays in C. elegans

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  • Mohammed D. Aljohani

    (King Abdullah University of Science and Technology (KAUST), Biological and Environmental Science and Engineering Division (BESE), KAUST Environmental Epigenetics Program (KEEP))

  • Sonia El Mouridi

    (King Abdullah University of Science and Technology (KAUST), Biological and Environmental Science and Engineering Division (BESE), KAUST Environmental Epigenetics Program (KEEP))

  • Monika Priyadarshini

    (King Abdullah University of Science and Technology (KAUST), Biological and Environmental Science and Engineering Division (BESE), KAUST Environmental Epigenetics Program (KEEP))

  • Amhed M. Vargas-Velazquez

    (King Abdullah University of Science and Technology (KAUST), Biological and Environmental Science and Engineering Division (BESE), KAUST Environmental Epigenetics Program (KEEP))

  • Christian Frøkjær-Jensen

    (King Abdullah University of Science and Technology (KAUST), Biological and Environmental Science and Engineering Division (BESE), KAUST Environmental Epigenetics Program (KEEP))

Abstract

Transgenes are prone to progressive silencing due to their structure, copy number, and genomic location. In C. elegans, repressive mechanisms are particularly strong in the germline with almost fully penetrant transgene silencing in simple extrachromosomal arrays and frequent silencing of single-copy transgene insertions. A class of non-coding DNA, Periodic An/Tn Clusters (PATCs) can prevent transgene-silencing in repressive chromatin or from small interfering RNAs (piRNAs). Here, we describe design rules (codon-optimization, intron and PATC inclusion, elevated temperature (25 °C), and vector backbone removal) for efficient germline expression from arrays in wildtype animals. We generate web-based tools to analyze PATCs and reagents for the convenient assembly of PATC-rich transgenes. An extensive collection of silencing resistant fluorescent proteins (e.g., gfp, mCherry, and tagBFP) can be used for dissecting germline regulatory elements and a set of enhanced enzymes (Mos1 transposase, Cas9, Cre, and Flp recombinases) enable efficient genetic engineering in C. elegans.

Suggested Citation

  • Mohammed D. Aljohani & Sonia El Mouridi & Monika Priyadarshini & Amhed M. Vargas-Velazquez & Christian Frøkjær-Jensen, 2020. "Engineering rules that minimize germline silencing of transgenes in simple extrachromosomal arrays in C. elegans," Nature Communications, Nature, vol. 11(1), pages 1-14, December.
    • Handle: RePEc:nat:natcom:v:11:y:2020:i:1:d:10.1038_s41467-020-19898-0
    DOI: 10.1038/s41467-020-19898-0
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    Cited by:

    1. Meghan Lee Arnold & Jason Cooper & Rebecca Androwski & Sohil Ardeshna & Ilija Melentijevic & Joelle Smart & Ryan J. Guasp & Ken C. Q. Nguyen & Ge Bai & David H. Hall & Barth D. Grant & Monica Driscoll, 2023. "Intermediate filaments associate with aggresome-like structures in proteostressed C. elegans neurons and influence large vesicle extrusions as exophers," Nature Communications, Nature, vol. 14(1), pages 1-17, December.
    2. Ichiro Aoki & Luca Golinelli & Eva Dunkel & Shripriya Bhat & Erschad Bassam & Isabel Beets & Alexander Gottschalk, 2024. "Hierarchical regulation of functionally antagonistic neuropeptides expressed in a single neuron pair," Nature Communications, Nature, vol. 15(1), pages 1-19, December.

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