| RRC ID |
66881
|
| 著者 |
You Y, Ramachandra SG, Jin T.
|
| タイトル |
A CRISPR-based method for testing the essentiality of a gene.
|
| ジャーナル |
Sci Rep
|
| Abstract |
The CRISPR/Cas9 system is a powerful method of editing genes by randomly introducing errors into the target sites. Here, we describe a CRISPR-based test for gene essentiality (CRISPR-E test) that allows the identification of essential genes. Specifically, we use sgRNA-mediated CRISPR/Cas9 to target the open reading frame of a gene in the genome and analyze the in-frame (3n) and frameshift (3n + 1 and 3n + 2) mutations in the targeted region of the gene in surviving cells. If the gene is non-essential, the cells would carry both in-frame (3n) and frameshift (3n + 1 and 3n + 2) mutations. In contrast, the cells would carry only in-frame (3n) mutations if the targeted gene is essential, and this selective elimination of frameshift (3n + 1 and 3n + 2) mutations of the gene indicate its essentiality. As a proof of concept, we have used this CRISPR-E test in the model organism Dictyostelium discoideum to demonstrate that Dync1li1 is an essential gene while KIF1A and fAR1 are not. We further propose a simple method for quantifying the essentiality of a gene using the CRISPR-E test.
|
| 巻・号 |
10(1)
|
| ページ |
14779
|
| 公開日 |
2020-9-8
|
| DOI |
10.1038/s41598-020-71690-8
|
| PII |
10.1038/s41598-020-71690-8
|
| PMID |
32901070
|
| PMC |
PMC7478968
|
| MeSH |
Amino Acid Sequence
Base Sequence
CRISPR-Cas Systems*
Dictyostelium / genetics*
Dictyostelium / growth & development
Dictyostelium / metabolism
Gene Editing*
Genes, Essential*
Protozoan Proteins / antagonists & inhibitors
Protozoan Proteins / genetics*
Protozoan Proteins / metabolism
Sequence Homology
|
| IF |
3.998
|
| リソース情報 |
| 細胞性粘菌 |
pTM1285 |
