Chapter title |
Building Cre Knockin Rat Lines Using CRISPR/Cas9
|
---|---|
Chapter number | 3 |
Book title |
Site-Specific Recombinases
|
Published in |
Methods in molecular biology, January 2017
|
DOI | 10.1007/978-1-4939-7169-5_3 |
Pubmed ID | |
Book ISBNs |
978-1-4939-7167-1, 978-1-4939-7169-5
|
Authors |
Yuanwu Ma, Lianfeng Zhang, Xingxu Huang |
Abstract |
Conditional gene inactivation strategy helps researchers to study the gene functions that are critical in embryogenesis or in defined tissues of adulthood. The Cre/loxP system is widely used for conditional gene inactivation/activation in cells or organisms. Cre knockin animal lines are essential for gene expression or inactivation in a spatially and temporally restricted manner. However, to generate a Cre knockin line by traditional approach is laborious. Recently, the clustered regularly interspaced short palindromic repeats and CRISPR-associated protein 9 (CRISPR/Cas9) has been proven as a simple and efficient genome-editing tool. We have used CRISPR/Cas9 system to generate rat strains that carry Cre genes in different targeted gene loci by direct delivery of gRNAs/Cas9/donors into fertilized eggs. Here, we described a stepwise procedure for the generation of Cre knockin rat, including target site selection, RNA preparation, the construction of the template donor, pronuclear injection, and the genotyping of precise Cre insertion in F0 rats. Taken together, the establishment of Cre knockin line can be achieved within 6 weeks. |
X Demographics
Geographical breakdown
Country | Count | As % |
---|---|---|
Unknown | 2 | 100% |
Demographic breakdown
Type | Count | As % |
---|---|---|
Members of the public | 2 | 100% |
Mendeley readers
Geographical breakdown
Country | Count | As % |
---|---|---|
Unknown | 15 | 100% |
Demographic breakdown
Readers by professional status | Count | As % |
---|---|---|
Student > Bachelor | 3 | 20% |
Student > Ph. D. Student | 3 | 20% |
Researcher | 2 | 13% |
Professor | 1 | 7% |
Other | 1 | 7% |
Other | 2 | 13% |
Unknown | 3 | 20% |
Readers by discipline | Count | As % |
---|---|---|
Agricultural and Biological Sciences | 4 | 27% |
Biochemistry, Genetics and Molecular Biology | 4 | 27% |
Neuroscience | 2 | 13% |
Pharmacology, Toxicology and Pharmaceutical Science | 1 | 7% |
Engineering | 1 | 7% |
Other | 0 | 0% |
Unknown | 3 | 20% |