Molecular genetics of bovine 70 kilodalton heat shock protein (HSP70) genes

Abstract

Sequences homologous to a 2.3 kilobase (kb) human HSP70 cDNA were extracted from a bovine sperm DNA library. The cloned DNA was restriction mapped and sequences containing HSP70 coding regions, along with those sequences containing unique, flanking DNA, were subcloned into plasmid vectors. Unique, flanking DNA fragments were used as locus-specific probes to identify the chromosomal location of the bovine HSP70 genes. DNA fragments containing HSP70 coding regions were sequenced using the dideoxy DNA sequencing procedure. Twenty-one positive clones were obtained and isolated for analysis. Based on restriction map data, the 21 clones can be categorized into 12 identical clone classes. Consolidating overlapping classes yields 3 regions of DNA containing 4 bovine HSP70 genes. Locus-specific probes were hybridized to bovine-rodent somatic cell hybrid panels to identify the chromosomal locations of the 3 regions of DNA containing HSP70 sequences. One region contains 2 tandemly arrayed genes (HSP70-1 and HSP70-2), and exists on chromosome 23, syntenic with BoLA, the bovine major histocompatibility complex (MHC). Single HSP70 sequences, designated HSP70-3 and HSP70-4, were found in two other genomic regions. Analysis of restriction enzyme digested bovine DNA has revealed at least 4 HSP70 restriction fragment length polymorphisms (RFLPs). By hybridizing the locus-specific probes to restriction enzyme digested bovine DNA and other methods, individual RFLPs have been assigned to the HSP70 genes isolated from the library. By combining three RFLPs which exist within a stretch of 20 kb (syntenic with bovine major histocompatibility complex), a highly polymorphic haplotype can be determined for each animal. This haplotype can be used as a marker in future linkage analyses involving the structure and gene order of the bovine MHC, as well as other loci on chromosome 23. Data obtained from DNA sequencing indicates that the four HSP70 sequences are highly conserved at both the nucleotide and presumed amino acid level. The addition of these loci to the bovine gene map further defines three regions of syntenic conservation between bovine and human genomes.