title: Specific cloning of DNA fragments absent from the DNA of a male patient with an X chromosome deletion.
abstract: A method that allows the specific cloning of DNA fragments absent from patients homozygous or hemizygous for chromosomal deletions is described. The method involves phenol-accelerated competitive DNA reassociation and subsequent molecular cloning of appropriately reassociated molecules. The deletion DNA sample utilized in the competition was isolated from a patient with a minute interstitial deletion in the short arm of the X chromosome. Sheared DNA isolated from a male child, who was diagnosed as having Duchenne muscular dystrophy, chronic granulomatous disease, and retinitis pigmentosa, was combined in a 200-fold excess with Mbo I-cleaved DNA isolated from a 49, XXXXY human lymphoid cell line, and the mixture was subjected to a phenol-enhanced reassociation technique. Analysis of 81 unique segments derived from cloned reassociated DNA molecules has led to the identification of 4 (5%) human DNA fragments that are absent from the male patient's DNA. The 4 clones were localized, on the basis of hybridization with restriction nuclease-digested genomic DNA from a panel of human and human-rodent hybrid cell lines, into three regions surrounding band 21 of the short arm of the normal human X chromosome. These clones are potential linkage markers for the diseases affecting this boy. Each clone, as well as others obtainable by this approach, may also serve as a starting point in the eventual cloning of these three X-linked-disease loci. Extension of this approach to other loci, including human tumors potentially homozygous for small deletions, should also be possible.
authors.forename: L M
authors.initials: LM
authors.lastname: Kunkel
authors.forename: A P
authors.initials: AP
authors.lastname: Monaco
authors.forename: W
authors.initials: W
authors.lastname: Middlesworth
authors.forename: H D
authors.initials: HD
authors.lastname: Ochs
authors.forename: S A
authors.initials: SA
authors.lastname: Latt
mesh_terms: D000818:Animals; D002460:Cell Line; D002872:Chromosome Deletion; D003001:Cloning, Molecular; D004247:DNA / Q000032:analysis*; D004262:DNA Restriction Enzymes; D005260:Female; D006801:Humans; D006822:Hybrid Cells / Q000166:cytology; D007621:Karyotyping; D008297:Male; D009693:Nucleic Acid Hybridization; D009695:Nucleic Acid Renaturation; D012729:Sex Chromosome Aberrations; D014960:X Chromosome
chemical_list: D004247:DNA; D004262:DNA Restriction Enzymes