Enrichment of cells with TALEN-induced mutations using surrogate reporters.
Targeted gene knockout using engineered nucleases such as transcription activator like-effector nucleases (TALENs) is a gold standard for investigating the functions of a gene of interest. Although most TALENs can cleave chromosomal DNA efficiently, the activities of designed TALENs are not always high enough to allow the efficient derivation of cells containing TALEN-driven mutations. Thus, simple methods to enrich cells containing TALEN-directed mutations would facilitate the use of TALENs. Here we describe the enrichment of such cells using surrogate episomal reporters coupled with flow cytometric sorting, magnetic separation, or hygromycin selection.
Graduate School of Biomedical Science and Engineering/College of Medicine, Hanyang University, Seoul, South Korea.
Young-Hoon
YH
Kim
Graduate School of Biomedical Science and Engineering/College of Medicine, Hanyang University, Seoul, South Korea.
Suresh
S
Ramakrishna
Graduate School of Biomedical Science and Engineering/College of Medicine, Hanyang University, Seoul, South Korea. Electronic address: hkim1@hanyang.ac.kr.
Hyongbum
H
Kim
Center for Genome Engineering, Institute for Basic Science, Seoul, South Korea; Department of Chemistry, Seoul National University, Seoul, South Korea. Electronic address: jskim01@snu.ac.kr.
Jin-Soo
JS
Kim
D001483:Base Sequence; D002469:Cell Separation / Q000379:methods*; D002934:Cinnamates; D003851:Deoxyribonucleases / Q000737:chemistry / Q000235:genetics; D005434:Flow Cytometry / Q000379:methods*; D017930:Genes, Reporter; D057809:HEK293 Cells; D006801:Humans; D006921:Hygromycin B / Q000031:analogs & derivatives; D008280:Magnetics; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009154:Mutation
D002934:Cinnamates; D006921:Hygromycin B; C026273:hygromycin A; D003851:Deoxyribonucleases
Enrichment; Flow cytometry; Hygromycin B; Magnetic separation; Mutant cells; Transcription activator-like effector nucleases