Stable expression of selectable genes introduced into human hematopoietic stem cells by electric field-mediated DNA transfer.

AUTOR(ES)

Toneguzzo, F

RESUMO

The successful introduction of DNA into human bone marrow cells by electric field-mediated transfer was initially demonstrated by the detection of transient chloramphenicol acetyltransferase (acetyl-CoA:chloramphenicol O3-acetyltransferase, EC 2,3.1.28) activity in marrow cell extracts. To determine whether DNA was transferred into hematopoietic stem cells, human nucleated marrow cells were subjected to electroporation in the presence of a plasmid construct containing the bacterial genes conferring resistance to the neomycin analogue G418 (neo) and to mycophenolic acid (gpt). The growth of granulocyte/macrophage colonies in selective media, followed by hybridization analyses of resistant cells, established that DNA was transferred into human granulopoietic progenitor cells and was stably maintained and expressed in their differentiated progeny. Electroporation, therefore, offers the opportunity to transfer genes effectively into human hematopoietic stem cells and avoids some of the disadvantages associated with other methods of gene transfer.

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