RESUMO
Radiation hybrid panels are already available for genome mapping in human and mouse. In this study we have used two model organisms (chicken and zebrafish) to show that hybrid panels that contain a full complement of the donor genome can be generated by fusion to hamster cells. The quality of the resulting hybrids has been assessed using PCR and FISH. We confirmed the utility of our panels by establishing the percentage of donor DNA present in the hybrids. Our hybrid resources will allow inexpensive gene mapping and we expect that this technology can be transferred to many other species. Such successes are providing the basis for a new era of mapping tools, in the form of whole genome radiation hybrid panels, and are opening new possibilities for systematic genome analysis in the animal genetics community.
Assuntos
Células Híbridas/efeitos da radiação , Animais , Linhagem Celular , Galinhas , Mapeamento Cromossômico , Cricetinae , Hibridização in Situ Fluorescente , Reação em Cadeia da Polimerase , Peixe-ZebraRESUMO
Chromo-helicase-DNA binding 1 (CHD1) is a conserved protein with a putative role in chromatin architecture. Single homologues have been found in mouse, Drosophila and yeast. In birds the situation is different as they possess two homologues. One is known to be W-linked, we show the second, closely related gene is linked to the Z sex chromosome. The basic structure of the Z-linked gene is similar to the homologous genes, however, it does possess an additional, internal 88 amino acid hydrophilic domain, rich in glutamic acid and lysine. Studies on pairs of genes sex-linked in mammals suggests rapid divergence of DNA sequence and function. We suggest the DNA sequences of CHD-W and CHD-Z do not follow this pattern.
Assuntos
Proteínas Aviárias , Galinhas/genética , Proteínas de Ligação a DNA/genética , Ligação Genética , Cromossomos Sexuais/genética , Sequência de Aminoácidos , Animais , Sequência de Bases , Embrião de Galinha , Mapeamento Cromossômico , DNA Complementar/genética , Feminino , Hibridização in Situ Fluorescente , Masculino , Dados de Sequência Molecular , Homologia de Sequência de AminoácidosRESUMO
The direct sequencing of the human type II procollagen (COL2A1) gene from polymerase chain reaction (PCR)-amplified genomic DNA is described. Thirty-two regions of the COL2A1 gene were asymmetrically amplified with intron primers which were specifically chosen to amplify a region spanning 500 to 800 bp of sequence encoding one or more exons and their accompanying intervening sequences. Primers for dideoxynucleotide sequencing of the PCR products were then designed to provide complete exon sequence information and to insure that intron:exon splice junction sequence data would be obtained. Amplification and sequencing reactions were performed on an automated workstation to facilitate the handling of multiple DNA templates. The procedure allowed efficient sequencing of over 25,000 bp of each allele of the COL2A1 gene per diploid genome. We used this method for the comparative analyses of COL2A1 sequences in DNA isolated from the blood of 42 unrelated individuals and we identified 21 neutral sequence variants in the gene. The sequence variations were confirmed by independent assays, including restriction enzyme digestion. The sequence variants described here will be important for identifying haplotypes of the type II procollagen gene that will be useful in defining a genetic etiology for diseases of cartilaginous tissues.
