Split selectable markers.
Nat Commun
; 10(1): 4968, 2019 10 31.
Article
en En
| MEDLINE
| ID: mdl-31672965
ABSTRACT
Selectable markers are widely used in transgenesis and genome editing for selecting engineered cells with a desired genotype but the variety of markers is limited. Here we present split selectable markers that each allow for selection of multiple "unlinked" transgenes in the context of lentivirus-mediated transgenesis as well as CRISPR-Cas-mediated knock-ins. Split marker gene segments fused to protein splicing elements called "inteins" can be separately co-segregated with different transgenic vectors, and rejoin via protein trans-splicing to reconstitute a full-length marker protein in host cells receiving all intended vectors. Using a lentiviral system, we create and validate 2-split Hygromycin, Puromycin, Neomycin and Blasticidin resistance genes as well as mScarlet fluorescent proteins. By combining split points, we create 3- and 6-split Hygromycin resistance genes, demonstrating that higher-degree split markers can be generated by a "chaining" design. We adapt the split marker system for selecting biallelically engineered cells after CRISPR gene editing. Future engineering of split markers may allow selection of a higher number of genetic modifications in target cells.
Texto completo:
1
Colección:
01-internacional
Base de datos:
MEDLINE
Asunto principal:
Ingeniería Genética
/
Técnicas de Transferencia de Gen
/
Empalme de Proteína
/
Farmacorresistencia Bacteriana
/
Inteínas
/
Proteínas Luminiscentes
Límite:
Humans
Idioma:
En
Revista:
Nat Commun
Asunto de la revista:
BIOLOGIA
/
CIENCIA
Año:
2019
Tipo del documento:
Article
País de afiliación:
Estados Unidos