Localization of repetitive and unique DNA sequences neighbouring the rabbit β-globin gene

Abstract

DNA fragments containing the rabbit β-globin gene can be detected in restriction endonuclease digests of rabbit DNA, using filter hybridizations with a labelled β-globin cDNA plasmid as a probo for the structural gene. Analysis of the reannealing kinetics of these β-globin DNA fragments has been performed by denaturing rabbit DNA and renaturing to various C 0 t ‡ ‡ Abbreviations used: C 0t , initial DNA concentration (mol.1 −)× time (s); kb, bases × 10 −3; cDNA, complementary DNA; bp, base-pairs values, then determining by hydroxyapatite chromatography and subsequent filter hybridization whether a given β-globin DNA fragment has annealed with other DNA via a repetitive DNA tract contained in the fragment, or instead has remained completely singlestranded. This approach has revealed the presence of an intermediate repetitive DNA sequence about 0.6 to 2.4 kb from the 3′ end of the rabbit β-globin structural gene. The presence of this repetitive sequence was confirmed by filter hybridization analysis of rabbit DNA with various DNA fragments isolated from a cloned segment of rabbit genomic DNA containing the β-globin gene. A purified rabbit DNA fragment containing sequences 0.9 to 2.1 kb from the 3′ end of the structural gene was found to hybridize to many DNA fragments in restriction endonuclease digests of rabbit DNA. In situ hybridizations to rabbit metaphase chromosomes showed that copies of this repetitive sequence were present on several chromosomes, and were preferentially localized in centromeric and telomeric regions. In contrast, the remaining section of the rabbit β-globin gene region examined, extending 1.4 kb from the 5′ end of the β-globin gene to 0.9 kb from the 3′ end of the gene and including the intervening sequences within the structural gene, was shown to consist of single copy DNA.