Title |
CRISPR/Cas9-mediated gene knockin in the hydroid Hydractinia symbiolongicarpus
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Published in |
BMC Genomics, September 2018
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DOI | 10.1186/s12864-018-5032-z |
Pubmed ID | |
Authors |
Steven M. Sanders, Zhiwei Ma, Julia M. Hughes, Brooke M. Riscoe, Gregory A. Gibson, Alan M. Watson, Hakima Flici, Uri Frank, Christine E. Schnitzler, Andreas D. Baxevanis, Matthew L. Nicotra |
Abstract |
Hydractinia symbiolongicarpus, a colonial cnidarian, is a tractable model system for many cnidarian-specific and general biological questions. Until recently, tests of gene function in Hydractinia have relied on laborious forward genetic approaches, randomly integrated transgenes, or transient knockdown of mRNAs. Here, we report the use of CRISPR/Cas9 genome editing to generate targeted genomic insertions in H. symbiolonigcarpus. We used CRISPR/Cas9 to promote homologous recombination of two fluorescent reporters, eGFP and tdTomato, into the Eukaryotic elongation factor 1 alpha (Eef1a) locus. We demonstrate that the transgenes are expressed ubiquitously and are stable over two generations of breeding. We further demonstrate that CRISPR/Cas9 genome editing can be used to mark endogenous proteins with FLAG or StrepII-FLAG affinity tags to enable in vivo and ex vivo protein studies. This is the first account of CRISPR/Cas9 mediated knockins in Hydractinia and the first example of the germline transmission of a CRISPR/Cas9 inserted transgene in a cnidarian. The ability to precisely insert exogenous DNA into the Hydractinia genome will enable sophisticated genetic studies and further development of functional genomics tools in this understudied cnidarian model. |
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Geographical breakdown
Country | Count | As % |
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United States | 7 | 39% |
Japan | 1 | 6% |
France | 1 | 6% |
Unknown | 9 | 50% |
Demographic breakdown
Type | Count | As % |
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Scientists | 10 | 56% |
Members of the public | 6 | 33% |
Practitioners (doctors, other healthcare professionals) | 1 | 6% |
Unknown | 1 | 6% |
Mendeley readers
Geographical breakdown
Country | Count | As % |
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Unknown | 85 | 100% |
Demographic breakdown
Readers by professional status | Count | As % |
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Student > Ph. D. Student | 18 | 21% |
Researcher | 11 | 13% |
Student > Master | 10 | 12% |
Student > Bachelor | 7 | 8% |
Other | 3 | 4% |
Other | 7 | 8% |
Unknown | 29 | 34% |
Readers by discipline | Count | As % |
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Biochemistry, Genetics and Molecular Biology | 22 | 26% |
Agricultural and Biological Sciences | 19 | 22% |
Environmental Science | 5 | 6% |
Immunology and Microbiology | 3 | 4% |
Medicine and Dentistry | 2 | 2% |
Other | 4 | 5% |
Unknown | 30 | 35% |