This subproject is one of many research subprojects utilizing theresources provided by a Center grant funded by NIH/NCRR. The subproject andinvestigator (PI) may have received primary funding from another NIH source,and thus could be represented in other CRISP entries. The institution listed isfor the Center, which is not necessarily the institution for the investigator.To generate lentiviral particles with improved long term site dependent expression in primate ES cells and their derivatives and to test specific genetic markers for their role in self renewal.In order to efficiently deliver exogenous genes into human embryonic stem cells (hESCs), we've optimized lentiviral expression system: 1) we decreased the vector size to improve the cloning of various genes; 2) we developed different expression vectors with different promoters such as EF1a promoter and PGK promoter to allow different levels of transgene expression, along with different drug-selection markers such as neomycin phosphotransferase and puromycin-N-acetyl-transferase; 3) We optimized lentiviral packaging protocols, allowing us to obtain viral particles with consistent high titers. We have successfully produced 14 new lentiviral constructs using genes found to be essential for pluripotency in ES cells. We have also produced lentiviral particles from these 14 new constructs. Using these lentiviral particles, we were able to successfully reprogram differentiated skin cells back to ES-like cells. The new cells are called ips cells. This research used WNPRC Stem Cell Resources, IS services and federally approved human ES cell lines.

Agency
National Institute of Health (NIH)
Institute
National Center for Research Resources (NCRR)
Type
Primate Research Center Grants (P51)
Project #
2P51RR000167-47A1
Application #
7716406
Study Section
Special Emphasis Panel (ZRR1-CM-8 (01))
Project Start
2008-07-23
Project End
2009-04-30
Budget Start
2008-07-23
Budget End
2009-04-30
Support Year
47
Fiscal Year
2008
Total Cost
$81,915
Indirect Cost
Name
University of Wisconsin Madison
Department
Type
Other Domestic Higher Education
DUNS #
161202122
City
Madison
State
WI
Country
United States
Zip Code
53715
Kang, HyunJun; Mesquitta, Walatta-Tseyon; Jung, Ho Sun et al. (2018) GATA2 Is Dispensable for Specification of Hemogenic Endothelium but Promotes Endothelial-to-Hematopoietic Transition. Stem Cell Reports 11:197-211
Rhoads, Timothy W; Burhans, Maggie S; Chen, Vincent B et al. (2018) Caloric Restriction Engages Hepatic RNA Processing Mechanisms in Rhesus Monkeys. Cell Metab 27:677-688.e5
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Park, Mi Ae; Jung, Ho Sun; Slukvin, Igor (2018) Genetic Engineering of Human Pluripotent Stem Cells Using PiggyBac Transposon System. Curr Protoc Stem Cell Biol 47:e63
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Shackman, A J; Fox, A S; Oler, J A et al. (2017) Heightened extended amygdala metabolism following threat characterizes the early phenotypic risk to develop anxiety-related psychopathology. Mol Psychiatry 22:724-732
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