Physical localization of two DNA markers closely linked to the cystic fibrosis locus by pulsed-field gel electrophoresis

Abstract

Our previous linkage analysis suggested that the DNA segment D7S122 is located between MET and D7S8, the two genetic markers that are thought to flank the cystic fibrosis locus (CF). Subsequent chromosome walking experiments revealed that D7S122 is within close distance to another randomly isolated DNA marker, D7S340. To determine the physical relationship among D7S122, D7S340, MET, and D7S8, we have constructed a long-range restriction map of the region containing these four DNA segments, by using DNA from a human/hamster somatic hybrid cell line 4AF-KO15 (containing a single human chromosome 7) and a series of rare-cutting restriction enzymes. The combined results of complete, partial, and double digestion analyses confirm that D7S122 and D7S340 are located between MET and D7S8. The order of these markers is MET-D7S340-D7S122-D7S8, with distance intervals of approximately 500, 10, and 980 kbp, respectively. Together with family analysis, this information will be useful for eventual identification of the CF gene.