The immediate goal of these studies is to develop methods for efficiently introducing human globin genes into hemopoietic cells both from tissue culture lines and normal bone marrow to study their tissue specific regulation. A hybrid SV 40 virus which contains the prokaryotic gene coding for chloramphenicol acetyl transferase (CAT) has been constructed and used to transfer, and transiently express the CAT gene in hemopoietic cell lines and fresh bone marrow cells of humans and other species. Recombinant SV 40 lysates contain wild type SV40 virus which severely compromises their use to introduce genes permanently into cells and cell lines. We have therefore made use of helper free recombinant adenoviruses that contain the neomycin resistant gene (neo) and have successfully transformed the hemopoietic cell lines K562 and MEL to G418 resistance. We have also constructed double recombinant adenoviruses containing the neo gene and the human beta globin gene. This virus can successfully transfer both genes into K562 and other cell lines. Although the transformation frequency of adenoviral vectors is considerably higher than that of DNA mediated transfer, it has not proven to be high enough for hemopoietic stem cells. Therefore retroviral vectors containing a human globin gene and a selectable marker gene are being constructed to achieve gene transfer into hematopoietic stem cells in vitro and in vivo.
