FACS-Assisted CRISPR-Cas9 Genome Editing Facilitates Parkinson's Disease Modeling.
Stem Cell Reports
; 9(5): 1423-1431, 2017 11 14.
Article
in En
| MEDLINE
| ID: mdl-28988985
ABSTRACT
Genome editing and human induced pluripotent stem cells hold great promise for the development of isogenic disease models and the correction of disease-associated mutations for isogenic tissue therapy. CRISPR-Cas9 has emerged as a versatile and simple tool for engineering human cells for such purposes. However, the current protocols to derive genome-edited lines require the screening of a great number of clones to obtain one free of random integration or on-locus non-homologous end joining (NHEJ)-containing alleles. Here, we describe an efficient method to derive biallelic genome-edited populations by the use of fluorescent markers. We call this technique FACS-assisted CRISPR-Cas9 editing (FACE). FACE allows the derivation of correctly edited polyclones carrying a positive selection fluorescent module and the exclusion of non-edited, random integrations and on-target allele NHEJ-containing cells. We derived a set of isogenic lines containing Parkinson's-disease-associated mutations in α-synuclein and present their comparative phenotypes.
Key words
Full text:
1
Database:
MEDLINE
Main subject:
Parkinson Disease
/
Alpha-Synuclein
/
Induced Pluripotent Stem Cells
/
CRISPR-Cas Systems
/
Gene Editing
Type of study:
Guideline
Limits:
Humans
Language:
En
Year:
2017
Type:
Article