Regulated expression of amplified human beta globin genes
D Rund, C Dobkin and A Bank
Gene therapy for the beta thalassemias and sickle cell anemia will require
high levels of expression of human beta globin genes. One method to achieve
this goal is amplification of globin genes transferred into the stem cells
in the bone marrow of these patients. If the amplified genes remain
normally regulated, they will then further increase their expression on
being induced to differentiate along an erythroid pathway. To begin this
study, we constructed a plasmid containing a neomycin resistance gene, a
human beta globin gene, and a wild-type DHFR cDNA, and transfected it into
mouse erythroleukemia cells. All the G418-resistant clones analyzed
acquired and expressed the human beta globin gene. By serial passage of the
cells in increasing concentrations of methotrexate, the exogenous human
beta globin genes were stably amplified in all lines, and all increased
their globin mRNA expression roughly proportional to their augmented copy
number. Most of the clones further increased their beta globin expression
on addition of an erythroid stimulus (dimethylsulfoxide). These results
indicate that globin gene amplification may be useful in increasing globin
mRNA expression in further experiments whose goal is gene therapy.
Volume 70,
Issue 3,
pp. 733-739,
09/01/1987
Copyright © 1987 by The American Society of Hematology
