CONSTRUCTION OF A SWINE YAC LIBRARY ALLOWING AN EFFICIENT RECOVERY OFUNIQUE AND CENTROMERIC REPEATED SEQUENCES

A swine DNA genomic library was constructed in yeast artificial chromo some (YAC) using the pYAC4 vector- and the AB1380 strain. The DNA prep ared from two Large White tnales was partially digested with EcoRI and size selected after both digestion and ligation. The YAC library cont ained 33792 arrayed clones with an average size of 280 kb as estimated by analysis of 2% of the clones, thus representing a threefold covera ge of the swine haploid genome. The library was organized in pools to facilitate the PCR screening. The complexity of the library was tested both for unique and centromeric repeated sequences. In all, 20 out of 22 primer sets allowed the characterization of one to six clones cont aining specific unique sequences. These sequences are known to be on C hromosomes (Chrs) 1, 2, 5, 6, 7, 8, 13, 14, 15, 17, and X. Eight addit ional clones carrying centromeric repeat units were also isolated with a single primer set. The sequencing of 37 distinct repeat units of ab out 340 bp subcloned from these eight YACs revealed high sequence dive rsity indicating the existence of numerous centromeric repent unit sub families in swine. Further-more, the analysis of the restriction patte rns with selected enzymes suggested a higher order organization of the repeat units. According to preliminary FISH experiments on a small nu mber of randomly chosen YACs and YACs carrying specific sequences, the chimerism appeared to be low. In addition, primed in situ labeling ex periments favored the idea that the YACs with centromeric repeat seque nces were derived from a subset of metacentric and submetacentric chro mosomes.