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 introduce a selectable marker such as neomycin resistance or EGFP by homologous recombination into a tissue specific gene.We identified culture conditions that allow human ES cells to efficiently differentiate into mesodermal lineages in defined conditions. Additionally we created a DNA vector that would tag, the brachyury gene with CD4, neomycin, and a soft tag sequence using homologous recombination. The successful tagging of the brachyury gene would allow us to purify early mesodermal cells with simple magnetic cell sorting technique. It would also enable us to study the functions of T gene in definite mesoderm commitment during human ES cell differentiation using a highly specific Soft tag antibody. The successful applications of Soft tag would also provide a universal tagging strategy to study the functions of many other genes, especially those lacking good antibodies. This research uses WNPRC Stem Cell Resources, the IS Division, and federally approved human ES cell lines.
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