The isolation and characterization of linked δ- and β-globin genes from a cloned library of human DNA

Abstract

A cloned library of large, random embryonic human DNA fragments was constructed and screened for β-globin sequences using the cloned human β-globin cDNA plasmid pJW102 (Wilson et al., 1978) as a hybridization probe. Two independent clones were obtained and then characterized by restriction endonuclease cleavage analysis, hybridization experiments and partial DNA sequencing. Each of the clones carries both the adult δ- and β-globin genes. The two genes are separated by approximately 5.4 kilobases (kb) of DNA and their orientation with respect to the direction of transcription is 5′-δ-β-3′. Both the δ-and β-globin genes contain a large noncoding intervening sequence (950 and 900 bp, respectively) located between the codons for amino acids 104 (arginine) and 105 (leucine). Although the location of the large intervening sequence within the coding regions of the two genes is identical, the two noncoding sequences bear little sequence homology. A second, smaller intervening sequence similar to that found in other mammalian β-globin genes was detected near the 5′ end of the human β-globin gene. The two independently isolated β-globin clones differ from each other by the presence of a Pst I restriction enzyme cleavage site within the large intervening sequence of the δ-globin gene of one of the clones. This suggests that the human DNA carried in the two clones was derived from two homologous chromosomes which were heterozygous for the Pst I restriction enzyme recognition sequence.