RESUMO
The ability to reliably express fluorescent reporters or other genes of interest is important for using human pluripotent stem cells (hPSCs) as a platform for investigating cell fates and gene function. We describe a simple expression system, designated GAPTrap (GT), in which reporter genes, including GFP, mCherry, mTagBFP2, luc2, Gluc, and lacZ are inserted into the GAPDH locus in hPSCs. Independent clones harboring variations of the GT vectors expressed remarkably consistent levels of the reporter gene. Differentiation experiments showed that reporter expression was reliably maintained in hematopoietic cells, cardiac mesoderm, definitive endoderm, and ventral midbrain dopaminergic neurons. Similarly, analysis of teratomas derived from GT-lacZ hPSCs showed that ß-galactosidase expression was maintained in a spectrum of cell types representing derivatives of the three germ layers. Thus, the GAPTrap vectors represent a robust and straightforward tagging system that enables indelible labeling of PSCs and their differentiated derivatives.
Assuntos
Expressão Gênica , Genes Reporter , Vetores Genéticos/genética , Células-Tronco Pluripotentes/metabolismo , Transgenes , Sistemas CRISPR-Cas , Diferenciação Celular , Linhagem Celular , Repetições Palindrômicas Curtas Agrupadas e Regularmente Espaçadas , Humanos , Células-Tronco Pluripotentes/citologia , Nucleases dos Efetores Semelhantes a Ativadores de TranscriçãoRESUMO
Klippel-Trenaunay syndrome (KTS) consists of a vascular nevus involving an extremity, varicosities of that extremity, and hypertrophy of bone and soft tissue. When arteriovenous malformation is also present, it is called Klippel-Trenaunay-Weber syndrome (KTWS). Ophthalmic features of these syndromes include vascular anomalies of the orbit, iris, retina, choroid, and optic nerve. We report a case of a 16-year-old girl with KTS who was found to have bilateral optic nerve and chiasmal gliomas, optic disk drusen, and acquired myelination of the retinal nerve fiber layer. These findings have not been previously reported to be associated with KTS or KTWS.