Search

The SOX gene family consists of a large number of embryonically expressed genes capable of encoding putative transcription factors and related by a DNA-binding domain, the HMG-box. We cloned and characterized the ovine SOX2 transcript using the screening of a testis (12dpp) cDNA library with a probe containing the SRY-HMG-box and we performed 3'RACE experiments. The ovine SOX2 sequence is strongly conserved in comparison to the human, mouse and chicken homologues and is located on sheep chromosome 1q33. The SOX2 expression pattern in developing gonads is consistent with the hypothesis that this gene plays a role in the germ cell line.

The ability to transfer genes into cells from different species with murine recombinant retroviruses was evaluated with the SVnls LacZ reporter gene. Mouse and cat packaging cell lines can be used to transfer amphotropic pseudotype, in human, mouse, cat, rabbit, sheep, horse and beef cells and with a very low efficiency in pig and avian cells. Xenotropic pseudotype recombinant retroviruses, produced in cat and rabbit packaging cell lines, transferred genes with the same efficiency as amphotropic retroviruses in human, cat, rabbit and sheep cells. In contrast to amphotropic retroviruses, xenotropic retroviruses infect beef, pig and horse cells with a high efficiency. These results emphasize the need to determine carefully the producer cell line (the type of helper virus and the species origin of the cell) for efficient transfer of genes in cells and embryos.

To obtain bovine Y-derived probes, we have constructed a bovine plasmid library enriched for Y-specific DNA sequences by the deletion enrichment method. The resulting clones were analyzed by hybridization to Southern blots of male and female genomic DNA. From 200 clones tested, two (BC1.2 and BC1.34) were entirely male specific, six gave a male-female differential hybridization pattern, and the remaining reacted similarly with male and female DNA. Interspecies somatic cell hybrid studies and chromosomal in situ hybridization confirmed that the BC1.2 sequence was derived from the Y chromosome. This 54-bp fragment is present at about 2000-2500 copies in the bovine male genome. No polymorphism was revealed with any of the restriction enzymes used, suggesting enzyme site conservation within blocks of repeats. Evolutionary study has shown that the BC1.2 sequence is conserved within Bos and Bison genera and remains male specific. The male specificity and repeated nature of the BC1.2 sequence have enabled us to use it as a molecular probe for sex determination on small numbers of cells by in situ hybridization.