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| Content Provider | World Health Organization (WHO)-Global Index Medicus |
|---|---|
| Author | Housden, Benjamin E. Hu, Yanhui Perrimon, Norbert |
| Description | Author Affiliation: Housden BE ( Department of Genetics, Harvard Medical School, Boston, Massachusetts 02115); Hu Y ( Department of Genetics, Harvard Medical School, Boston, Massachusetts 02115); Perrimon N ( Department of Genetics, Harvard Medical School, Boston, Massachusetts 02115) |
| Abstract | The recent advances in CRISPR-based genome engineering have enabled a plethora of new experiments to study a wide range of biological questions. The major attraction of this system over previous methods is its high efficiency and simplicity of use. For example, whereas previous genome engineering technologies required the generation of new proteins to target each new locus, CRISPR requires only the expression of a different single guide RNA (sgRNA). This sgRNA binds to the Cas9 endonuclease protein and directs the generation of a double-strand break to a highly specific genomic site determined by the sgRNA sequence. In addition, the relative simplicity of the Drosophila genome is a particular advantage, as possible sgRNA off-target sites can easily be avoided. Here, we provide a step-by-step protocol for designing sgRNA target sites using the Drosophila RNAi Screening Center (DRSC) Find CRISPRs tool (version 2). We also describe the generation of sgRNA expression plasmids for the use in cultured Drosophila cells or in vivo. Finally, we discuss specific design requirements for various genome engineering applications. |
| File Format | HTM / HTML |
| ISSN | 19403402 |
| Issue Number | 9 |
| Journal | Cold Spring Harbor Protocols |
| Volume Number | 2016 |
| e-ISSN | 15596095 |
| Language | English |
| Publisher | Cold Spring Harbor Laboratory Press |
| Publisher Date | 2016-09-01 |
| Publisher Place | United States |
| Access Restriction | One Nation One Subscription (ONOS) |
| Subject Keyword | Discipline Clinical Laboratory Techniques |
| Content Type | Text |
| Resource Type | Article |
| Subject | Biochemistry, Genetics and Molecular Biology |
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