Gagan Bajwa

Publications (2)11.93 Total impact

• Article: Inducible cassette exchange: a rapid and efficient system enabling conditional gene expression in embryonic stem and primary cells.

  Michelina Iacovino, Darko Bosnakovski, Holger Fey, Danielle Rux, Gagan Bajwa, Elisabeth Mahen, Ana Mitanoska, Zhaohui Xu, Michael Kyba
  [show abstract] [hide abstract]
  ABSTRACT: Genetic modification is critically enabling for studies addressing specification and maintenance of cell fate; however, methods for engineering modifications are inefficient. We demonstrate a rapid and efficient recombination system in which an inducible, floxed cre allele replaces itself with an incoming transgene. We target this inducible cassette exchange (ICE) allele to the (HPRT) locus and demonstrate recombination in murine embryonic stem cells (ESCs) and primary cells from derivative ICE mice. Using lentivectors, we demonstrate recombination at a randomly integrated ICE locus in human ESCs. To illustrate the utility of this system, we insert the myogenic regulator, Myf5, into the ICE locus in each platform. This enables efficient directed differentiation of mouse and human ESCs into skeletal muscle and conditional myogenic transdetermination of primary cells cultured in vitro. This versatile tool is thus well suited to gain-of-function studies probing gene function in the specification and reprogramming of cell fate.
  Stem Cells 10/2011; 29(10):1580-8. · 7.78 Impact Factor
• Article: A conserved role for Hox paralog group 4 in regulation of hematopoietic progenitors.

  Michelina Iacovino, Carmen Hernandez, Zhaohui Xu, Gagan Bajwa, Melissa Prather, Michael Kyba
  [show abstract] [hide abstract]
  ABSTRACT: Regulatory circuits that control stem cell fate decisions can be identified and understood by manipulating individual regulatory elements genetically. While impractical in the rare somatic stem cells of primary tissue, this approach is feasible in embryonic stem cells differentiated in vitro into the somatic stem cell type of interest. We present an improved highly efficient targeting system allowing genes to be integrated into a predetermined, doxycycline-inducible locus, and corresponding inducible embryonic stem cell lines to be generated rapidly. We apply this system to evaluate a key hematopoietic progenitor cell regulatory element, HoxB4, and its mammalian paralogs, whose effects have not yet been tested in this context. We show that all Hox paralog group 4 members, A4, B4, C4, and D4, have similar effects on hematopoietic stem and progenitor self-renewal in vitro, and thus classify Hox paralog group 4 as promoting self-renewal. Each paralog group 4 member both promotes proliferation and inhibits differentiation, enabling the exponential expansion of hematopoietic progenitors from the c-kit(+)/CD41(+) cell fraction of day 6 murine embryoid bodies. By evaluating a set of deletion mutants we show that sequences in addition to the homeodomain and hexapeptide motif are required for this activity. These results highlight the utility of this expression system to perform functional and structural analyses of genetic regulators of cell fate decisions.
  Stem cells and development 10/2008; 18(5):783-92. · 4.15 Impact Factor