COMPARATIVE ZOO-FISH ANALYSIS WITH RIVER BUFFALO 1Q PAINTING PROBE REVEALED THE CONSERVATION OF SUCH CHROMOSOME IN ALPACA (Vicugna pacos)

The alpaca (Vicugna pacos, 2n=74) belongs to the family Camelidae, order Cetartiodactyla. Karyotypic studies indicated extensive similarities within all members of the family with only slight variations in the amount and distribution of heterochromatin. Recently, molecular cytogenetic investigation led to the establishment of genome-wide comparative maps between camel, cattle, pig and human. Cross-species chromosome painting represents a very useful approach to define homologous DNA conserved segments. In this study we perform a comparative ZOO-FISH with river buffalo (BBU, 2n=50) 1q probe to identify the homologous chromosomes in alpaca. BBU1q painting probe was prepared through microdissection and DOPPCR. Briefly, fixed lymphocytes were spread onto a 24x60 mm coverslip, which was treated for GTG-banding. 15 copies of the 1q chromosome were scraped from the coverslip, collected and amplified by DOP-PCR. A second PCR reaction was performed for the labeling of DNA with digoxigenin-11-dUTP. Fluorescent in situ hybridization was carried out according to standard protocols. The labeled probe was revealed by antidig-rhodamine. Slides were counterstained with DAPI in antifade. The painting probe produced by chromosome microdissection and DOPPCR was tested first on river buffalo metaphases to verify the specific hybridization on the long arm of BBU1q. Then, the same probe was crosshybridized on alpaca chromosomes (VPA). A strong fluorescent signal was detected on VPA1q. According to the standard karyotype of alpaca and the DAPI banding, the signal covers the entire region of VPA1q1, whereas the region VPA1q2, including the telomere, did not show any signal. This result adds further interesting knowledge to the comparative cytogenetics of the farm animals and their evolution. Further buffalo painting probes will be cross-hybridized on alpaca metaphases to shed light on the comparative map of these two species. This study was supported by CISIA-VARIGEAV project, National Research Council (CNR) of Italy.