Sequence-specific modification of a beta-thalassemia locus by small DNA fragments in human erythroid progenitor cells.
Gene therapy has been proposed as a definitive cure of beta-thalassemia. We applied a gene targeting approach, based on the introduction of small DNA fragments (SDF) into erythroid progenitor cells, to specifically modify the beta-globin gene sequence at codon 39. The strategy was first tested in normal individuals by delivering mutant SDF that were able to produce the beta-39 (C->T) mutation. Secondly, wild-type SDF were electroporated into target cells of beta-3i9/beta-39 b-thalassemic patients to correct the endogenous mutation. In both cases, gene modification was assayed by allele-specific polymerase chain reaction of DNA and mRNA, by restriction fragment length polymorphism analysis and by direct sequencing.
Colosimo
Alessia
A
Guida
Valentina
V
Antonucci
Ivana
I
Bonfini
Tiziana
T
Stuppia
Liborio
L
Dallapiccola
Bruno
B
D000483:Alleles; D001483:Base Sequence; D003062:Codon; D053938:DNA Fragmentation; D018274:Electroporation; D015672:Erythroid Precursor Cells / Q000166:cytology*; D018390:Gene Targeting; D005914:Globins / Q000235:genetics; D006579:Heterozygote; D006801:Humans; D008969:Molecular Sequence Data; D009154:Mutation; D016133:Polymerase Chain Reaction; D012333:RNA, Messenger / Q000378:metabolism; D017086:beta-Thalassemia / Q000235:genetics*
D003062:Codon; D012333:RNA, Messenger; D005914:Globins