Title |
Single Cas9 nickase induced generation of NRAMP1 knockin cattle with reduced off-target effects
|
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Published in |
Genome Biology, February 2017
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DOI | 10.1186/s13059-016-1144-4 |
Pubmed ID | |
Authors |
Yuanpeng Gao, Haibo Wu, Yongsheng Wang, Xin Liu, Linlin Chen, Qian Li, Chenchen Cui, Xu Liu, Jingcheng Zhang, Yong Zhang |
Abstract |
The CRISPR-Cas9 system is a widely utilized platform for transgenic animal production in various species, although its off-target effects should be addressed. Several applications of this tool have been proposed in model animals but remain insufficient for transgenic livestock production. Here, we report the first application of single Cas9 nickase (Cas9n) to induce gene insertion at a selected locus in cattle. We identify the main binding sites of a catalytically inactive Cas9 (dCas9) protein in bovine fetal fibroblast cells (BFFs) with chromatin immunoprecipitation sequencing (ChIP-seq). Subsequently, we demonstrate that a single Cas9n-induced single-strand break can stimulate the insertion of the natural resistance-associated macrophage protein-1 (NRAMP1) gene with reduced, but still considerable, off-target effects. Through somatic cell nuclear transfer, we finally obtain transgenic cattle with increased resistance to tuberculosis. Our results contribute to the development of CRISPR-Cas9 system for agriculture applications. |
X Demographics
Geographical breakdown
Country | Count | As % |
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United Kingdom | 6 | 21% |
United States | 3 | 10% |
Australia | 3 | 10% |
Saudi Arabia | 2 | 7% |
Italy | 2 | 7% |
France | 1 | 3% |
Spain | 1 | 3% |
Chile | 1 | 3% |
Thailand | 1 | 3% |
Other | 3 | 10% |
Unknown | 6 | 21% |
Demographic breakdown
Type | Count | As % |
---|---|---|
Members of the public | 15 | 52% |
Scientists | 12 | 41% |
Science communicators (journalists, bloggers, editors) | 1 | 3% |
Practitioners (doctors, other healthcare professionals) | 1 | 3% |
Mendeley readers
Geographical breakdown
Country | Count | As % |
---|---|---|
Finland | 1 | <1% |
Sweden | 1 | <1% |
Unknown | 272 | 99% |
Demographic breakdown
Readers by professional status | Count | As % |
---|---|---|
Student > Bachelor | 52 | 19% |
Student > Master | 40 | 15% |
Researcher | 32 | 12% |
Student > Ph. D. Student | 28 | 10% |
Other | 11 | 4% |
Other | 36 | 13% |
Unknown | 75 | 27% |
Readers by discipline | Count | As % |
---|---|---|
Agricultural and Biological Sciences | 87 | 32% |
Biochemistry, Genetics and Molecular Biology | 70 | 26% |
Veterinary Science and Veterinary Medicine | 7 | 3% |
Chemistry | 5 | 2% |
Social Sciences | 4 | 1% |
Other | 20 | 7% |
Unknown | 81 | 30% |