Submitted on March 23, 2006
Accepted on September 14, 2006
Production of GFP transgenic embryonic stem cells using the GENSAT bacterial artificial chromosome library
Mark J Tomishima 1*,
Anna-Katerina Hadjantonakis 2,
Shiaoching Gong 3,
Lorenz Studer 1
1 Developmental Biology Program, Sloan-Kettering Institute, New York, New York; Division of Neurosurgery, Sloan-Kettering Institute, New York, New York
2 Developmental Biology Program, Sloan-Kettering Institute, New York, New York
3 Rockefeller University, New York, New York
* To whom correspondence should be addressed. E-mail: tomishim{at}mskcc.org.
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Abstract |
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Transgenic GFP reporter embryonic stem (ES) cells are powerful tools for studying gene regulation and lineage choice during development. Here we present a rapid method for the generation of ES cells expressing GFP under control of selected genes. Bacterial artificial chromosomes (BACs) from a previously constructed GFP transcriptional fusion library (GENSAT) were modified for use in ES cells, and multiple BAC transgenic ES cell lines were generated. Specific GFP expression in transgenic cell lines was confirmed during neural differentiation marking neural stem cells, neuronal precursors and glial progeny by Hes5, Dll1 and GFAP, respectively. GFP was dynamically regulated in ES cell progeny in response to soluble factors that inhibit Notch signaling and a factor that directs astroglial fate choice. Our protocols provide a simple and efficient strategy to utilize the whole GENSAT BAC library to create hundreds of novel fluorescent cell lines for use in ES cell biology.
