In vivo tracing of the Cytokeratin 14 lineages using self-cleaving guide RNAs and CRISPR/Cas9.
Dev Biol
; 504: 120-127, 2023 12.
Article
in En
| MEDLINE
| ID: mdl-37813160
The current gold-standard for genetic lineage tracing in transgenic mice is based on cell-type specific expression of Cre recombinase. As an alternative, we developed a cell-type specific CRISPR/spCas9 system for lineage tracing. This method relies on RNA polymerase II promoter driven self-cleaving guide RNAs (scgRNA) to achieve tissue-specificity. To demonstrate proof-of-principle for this approach a transgenic mouse was generated harbouring a knock-in of a scgRNA into the Cytokeratin 14 (Krt14) locus. Krt14 expression marks the stem cells of squamous epithelium in the skin and oral mucosa. The scgRNA targets a Stop cassette preceding a fluorescent reporter in the Ai9-tdtomato mouse. Ai9-tdtomato reporter mice harbouring this allele along with a spCas9 transgene demonstrated precise marking of the Krt14 lineage. We conclude that RNA polymerase II promoter driven scgRNAs enable the use of CRISPR/spCas9 for genetic lineage tracing.
Full text:
1
Collection:
01-internacional
Database:
MEDLINE
Main subject:
RNA Polymerase II
/
CRISPR-Cas Systems
Limits:
Animals
Language:
En
Journal:
Dev Biol
Year:
2023
Type:
Article