A homologous recombination (HR) approach for site-specific correction of mutations would be highly desirable for the treatment of genetic disorders if recombination efficiencies were sufficiently high as to permit a biological effect. Using a T cell thymoma line derived from severe combined immunodeficient (SCID) mice with a point mutation in the gene encoding the DNA-dependent protein kinase catalytic subunit (DNA-PKcs), we have shown that short DNA fragments (SDFs; 621 bases) can provide genotypic and functional correction of these cells. Double-stranded SDFs (dsSDFs) or single-stranded SDFs (ssSDFs) were designed to span the wild-type sequence of exon 85 in the DNA-PKcs gene and part of the 3' and 5' flanking intron regions. SCID cells were nucleofected with both single- and double-stranded wild-type SDF sequences. Corrected cells were selected on the basis of protection from radiation hypersensitivity that occurs as a consequence of the SCID mutation. Correction was mediated by both SDF forms (double and single stranded). These results indicate that SDFs can correct point mutations by HR with the possibility of harnessing ionizing radiation (IR) as a selection method to eliminate noncorrected cells and enrich for corrected SCID radioresistant cells.