GAPTrap: A Simple Expression System for Pluripotent Stem Cells and Their Derivatives.
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 β-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.
Murdoch Childrens Research Institute, Flemington Road, Parkville, VIC 3052, Australia; Department of Paediatrics, University of Melbourne, Parkville, VIC 3050, Australia.
Tim
T
Kao
Murdoch Childrens Research Institute, Flemington Road, Parkville, VIC 3052, Australia.
Tanya
T
Labonne
The Florey Institute of Neuroscience and Mental Health, Melbourne University, Parkville, VIC 3052, Australia.
Jonathan C
JC
Niclis
Department of Paediatrics, University of Melbourne, Parkville, VIC 3050, Australia.
Ritu
R
Chaurasia
Murdoch Childrens Research Institute, Flemington Road, Parkville, VIC 3052, Australia.
Zerina
Z
Lokmic
Murdoch Childrens Research Institute, Flemington Road, Parkville, VIC 3052, Australia; Department of Paediatrics, University of Melbourne, Parkville, VIC 3050, Australia.
Elizabeth
E
Qian
Murdoch Childrens Research Institute, Flemington Road, Parkville, VIC 3052, Australia; Department of Anatomy and Developmental Biology, Monash University, Clayton, VIC 3800, Australia.
Freya F
FF
Bruveris
Murdoch Childrens Research Institute, Flemington Road, Parkville, VIC 3052, Australia; Department of Paediatrics, University of Melbourne, Parkville, VIC 3050, Australia.
Sara E
SE
Howden
Murdoch Childrens Research Institute, Flemington Road, Parkville, VIC 3052, Australia; Department of Paediatrics, University of Melbourne, Parkville, VIC 3050, Australia.
Ali
A
Motazedian
Department of Anatomy and Developmental Biology, Monash University, Clayton, VIC 3800, Australia; Division of Developmental Biology, Cincinnati Children's Hospital Medical Centre, Cincinnati, OH 45229, USA.
Jacqueline V
JV
Schiesser
Australian Centre for Blood Diseases, Monash University, The Alfred Centre, Melbourne, VIC 3004, Australia.
Magdaline
M
Costa
Murdoch Childrens Research Institute, Flemington Road, Parkville, VIC 3052, Australia.
Koula
K
Sourris
Murdoch Childrens Research Institute, Flemington Road, Parkville, VIC 3052, Australia.
Elizabeth
E
Ng
Murdoch Childrens Research Institute, Flemington Road, Parkville, VIC 3052, Australia.
David
D
Anderson
Department of Anatomy and Developmental Biology, Monash University, Clayton, VIC 3800, Australia.
Antonietta
A
Giudice
Murdoch Childrens Research Institute, Flemington Road, Parkville, VIC 3052, Australia; Department of Paediatrics, University of Melbourne, Parkville, VIC 3050, Australia.
Peter
P
Farlie
Murdoch Childrens Research Institute, Flemington Road, Parkville, VIC 3052, Australia; Department of Paediatrics, University of Melbourne, Parkville, VIC 3050, Australia; Department of Cardiology, The Royal Children's Hospital, Parkville, VIC 3052, Australia.
Michael
M
Cheung
Murdoch Childrens Research Institute, Flemington Road, Parkville, VIC 3052, Australia; Department of Paediatrics, University of Melbourne, Parkville, VIC 3050, Australia.
Shireen R
SR
Lamande
Murdoch Childrens Research Institute, Flemington Road, Parkville, VIC 3052, Australia; Department of Paediatrics, University of Melbourne, Parkville, VIC 3050, Australia.
Anthony J
AJ
Penington
The Florey Institute of Neuroscience and Mental Health, Melbourne University, Parkville, VIC 3052, Australia.
Clare L
CL
Parish
The Florey Institute of Neuroscience and Mental Health, Melbourne University, Parkville, VIC 3052, Australia.
Lachlan H
LH
Thomson
Stem Cell and Microenvironment Laboratory, Weill Cornell Medical College in Qatar, Qatar Foundation, Education City, Doha, Qatar; Department of Genetic Medicine, Weill Cornell Medical College, New York, NY 10065-4896, USA.
Arash
A
Rafii
Murdoch Childrens Research Institute, Flemington Road, Parkville, VIC 3052, Australia; School of Biosciences, University of Melbourne, Parkville, VIC 3050, Australia.
David A
DA
Elliott
Murdoch Childrens Research Institute, Flemington Road, Parkville, VIC 3052, Australia; Department of Paediatrics, University of Melbourne, Parkville, VIC 3050, Australia; Department of Anatomy and Developmental Biology, Monash University, Clayton, VIC 3800, Australia. Electronic address: andrew.elefanty@mcri.edu.au.
Andrew G
AG
Elefanty
Murdoch Childrens Research Institute, Flemington Road, Parkville, VIC 3052, Australia; Department of Paediatrics, University of Melbourne, Parkville, VIC 3050, Australia; Department of Anatomy and Developmental Biology, Monash University, Clayton, VIC 3800, Australia. Electronic address: ed.stanley@mcri.edu.au.
Edouard G
EG
Stanley
D064113:CRISPR-Cas Systems; D002454:Cell Differentiation; D002460:Cell Line; D064112:Clustered Regularly Interspaced Short Palindromic Repeats; D015870:Gene Expression; D017930:Genes, Reporter; D005822:Genetic Vectors / Q000235:genetics*; D006801:Humans; D039904:Pluripotent Stem Cells / Q000166:cytology / Q000378:metabolism*; D000069896:Transcription Activator-Like Effector Nucleases; D019076:Transgenes
D000069896:Transcription Activator-Like Effector Nucleases
GAPDH; differentiation; expression system; human pluripotent stem cells; lineage tracing; reporter genes