A versatile tool for tracking the differentiation of human embryonic stem cells

The ability of human embryonic stem cells (hESCs) to undergo indefinite self-renewal in vitro and to produce lineages derived from all three embryonic germ layers both in vitro and in vivo makes such cells extremely valuable in both clinical and research settings. However, the generation of specialized cell lineages from a mixture of differentiated hESCs remains technically difficult. Tissue specific promoter-driven reporter genes are powerful tools for tracking cell types of interest in differentiated cell populations. Here, we describe the construction of modular lentivectors containing different tissue-specific promoters ( Tα1 of α-tubulin; aP2 of adipocyte Protein 2; and AFP of alpha fetoprotein) driving expression of humanized Renilla green fluorescent protein ( hrGFP ). To this end, we used MultiSite gateway technology and employed the novel vectors to successfully monitor hESC differentiation. We present a versatile method permitting target cells to be traced. Our system will facilitate research in developmental biology, transplantation, and in vivo stem cell tracking.