Open AccessHigh‐Level Sustained Transgene Expression in Human Embryonic Stem Cells Using Lentiviral Vectors
Author(s) -
Ma Yue,
Ramezani Ali,
Lewis Rachel,
Hawley Robert G.,
Thomson James A.
Publication year - 2003
Publication title -
stem cells
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 2.159
H-Index - 229
eISSN - 1549-4918
pISSN - 1066-5099
DOI - 10.1634/stemcells.21-1-111
Subject(s) - biology , transgene , vesicular stomatitis virus , viral vector , green fluorescent protein , stem cell , multiplicity of infection , embryonic stem cell , microbiology and biotechnology , genetic enhancement , locus control region , transduction (biophysics) , cell culture , gene expression , gene , genetics , enhancer , recombinant dna , biochemistry
Here we describe the sustained expression of transgenes introduced into human embryonic stem (ES) cells using self‐inactivating lentiviral vectors. At low multiplicity of infection, vesicular stomatitis virus‐pseudotyped vectors containing a green fluorescent protein (GFP) transgene under the control of a human elongation factor 1α promoter transduced human ES cells at high efficiency. The majority of the transduced ES cells, which harbored low numbers of integrated vectors, continued to express GFP after 60 days of culture. Incorporation of a scaffold attachment region (SAR) from the human interferon‐β gene into the lentiviral vector backbone increased the average level of GFP expression, and inclusion of the SAR together with a chromatin insulator from the 5′ end of the chicken β‐globin locus reduced the variability in GFP expression. When the transduced ES cells were induced to differentiate into CD34 + hematopoietic precursors in vitro, GFP expression was maintained with minimal silencing. The ability to efficiently introduce active transgenes into human ES cells will facilitate gain‐of‐function studies of early developmental processes in the human system. These results also have important implications for the possible future use of gene‐modified human ES cells in transplantation and tissue regeneration applications.