Strategic targeting of Cas9 nickase induces large segmental duplications.
Cell Genom
; 4(8): 100610, 2024 Aug 14.
Article
de En
| MEDLINE
| ID: mdl-39053455
ABSTRACT
Gene/segmental duplications play crucial roles in genome evolution and variation. Here, we introduce paired nicking-induced amplification (PNAmp) for their experimental induction. PNAmp strategically places two Cas9 nickases upstream and downstream of a replication origin on opposite strands. This configuration directs the sister replication forks initiated from the origin to break at the nicks, generating a pair of one-ended double-strand breaks. If homologous sequences flank the two break sites, then end resection converts them to single-stranded DNAs that readily anneal to drive duplication of the region bounded by the homologous sequences. PNAmp induces duplication of segments as large as â¼1 Mb with efficiencies exceeding 10% in the budding yeast Saccharomyces cerevisiae. Furthermore, appropriate splint DNAs allow PNAmp to duplicate/multiplicate even segments not bounded by homologous sequences. We also provide evidence for PNAmp in mammalian cells. Therefore, PNAmp provides a prototype method to induce structural variations by manipulating replication fork progression.
Texte intégral:
1
Collection:
01-internacional
Base de données:
MEDLINE
Sujet principal:
Saccharomyces cerevisiae
Limites:
Humans
Langue:
En
Journal:
Cell Genom
Année:
2024
Type de document:
Article
Pays d'affiliation:
Japon
Pays de publication:
États-Unis d'Amérique