Rapid detection of the highly polymorphic beta globin framework by denaturing gradient gel electrophoresis.

DGGI The detection and identification of point mutations responsible for common human genetic diseases still represent a major technical problem especially when analysing genes with a large coding region or a heterogeneous spectrum of mutations or both. Before the advent of the polymerase chain reaction (PCR) this problem could be partially circumvented by the identification of haplotypes of restriction fragment length polymorphisms (RFLPs) distributed around and within the gene of interest.' In informative families linkage between any given haplotype and the mutated gene can therefore be established, thus making the early and presymptomatic diagnosis of the disease feasible.'2 Several RFLPs distributed over more than 60 kb have been described in the 1 globin gene cluster.34 These polymorphisms have been observed to be non-randomly associated in different populations, thus giving rise to a limited number of haplotypes. Linkage disequilibrium has been shown between the haplotypes and 1 thalassaemia mutations, which make them extremely useful for the early diagnosis of the thalassaemia syndromes.2 Within the ,B globin gene itself, neutral substitutions have also been described. Extensive sequence analysis of normal and mutant ,B globin genes has indicated the existence of four normal P globin subhaplotypes, generally referred to as frameworks.45 These frameworks are particularly useful in linkage studies because they are located within the gene and are thus in tight linkage with the 13 thalassaemia mutations. The introduction of PCR has made the direct identification of single nucleotide substitutions, microdeletions, and insertions feasible. In particular the combination of PCR