242 DEVELOPMENT OF BAC FISH PROBES FOR GENETIC ANALYSIS OF NON-HUMAN PRIMATE GAMETES AND EMBRYONIC STEM CELLS
L. Froenicke A , S.M. Nichols B , H.M. Kubisch C , L.A. Lyons A , B.D. Bavister A D and C.A. Brenner A DA School of Veterinary Medicine, University of California Davis, Davis, CA 95616, USA
B Department of Biological Sciences, University of New Orleans, New Orleans, LA 70148, USA
C Divisions of Veterinary Medicine and Comparative Pathology, Tulane National Primate Research Center, Covington, LA 70433, USA
D Tulane Institute for Reproductive Medicine, Tulane University Medical School, New Orleans, LA 70112, USA. Email: cbrenner@uno.edu
Reproduction, Fertility and Development 17(2) 271-272 https://doi.org/10.1071/RDv17n2Ab242
Submitted: 1 August 2004 Accepted: 1 October 2004 Published: 1 January 2005
Abstract
Several factors relating to reproductive health and gamete competence must be considered when implementing programs for propagation of genetically valuable primates via natural breeding or assisted reproductive technologies. These relevant factors include ascertaining gamete chromosome normality of individuals. Our early attempts at characterizing aneuploidy levels of gametes within a model primate species, the rhesus macaque, utilizing Vysis centromeric fluorescent in situ hybridization (FISH) probes, failed to produce a signal. Commercially available probe sets for interphase cytogenetics rely partly on repetitive DNA probes. Therefore, we have employed either pools of human BACs or clones selected from a rhesus macaque BAC library for interphase cytogenetics. Bacterial artificial chromosome (BAC) FISH probes specific to regions of human DNA that are homologous to macaque DNA were produced, in collaboration with researchers at the University of California-Davis. The most common aneuploidies implicated in human birth defects or pregnancy failures are those involving the sex chromosomes and chromosomes HSA (human) 13, 16, 18, and 21 (Munne 2003 Placenta 24, 70–76). Hence, we have initially used human BAC FISH probes on the equivalent chromosome homologs in Macaca mulatta (MMU): X, Y, MMU 17 (HSA13), MMU 20 (HSA16), and MMU18 (HSA18). The homolog of HSA 21 is part of a larger chromosome in the macaque (MMU3) that also contains HSA 7 (based on nomenclature by Cambefort et al. 1976 Ann. Genet. 19, 5–9). Since aneuploidy has been linked to chromosome size and MMU3 is large, this macaque chromosome may be involved in aneuploidies. In addition, we determined the suitability of commercially available “chromosome paints” that target repetitive sequences and are widely used in determining karyotypes in human fertility clinics. Initial work was performed on cultured macaque blood cells exposed to Colcemid. Our preliminary data clearly demonstrate the feasibility of using probes that are commonly used in the analysis of human karyotypes to examine chromosomal anomalies in macaque gametes and embryos. Additional probes for other chromosomes are currently under investigation. The development of a large panel of available probes will provide a rapid and accurate method to assess aneuploidy/mosaicism levels in genetically valuable primate spermatozoa, in vitro-matured oocytes and blastomeres of IVP embryos. Furthermore, these probes will be useful to determine the karyotypic normalcy of cultured non-human primate embryonic stem cells.
This research has been supported by the University of New Orleans to CAB, Tulane National Primate Research Center base grant NIH 5P51 RR00164-41, and the Joe W. and Dorothy Dorsett Brown Foundation Cellular Imaging Facility.
