Production of pig-mink cell hybrids with single pig chromosomes 2, 5, 12, or t(1,13)

Monochromosomal cell hybrids are widely used in humans for chromosome mapping and for building chromosome-specific libraries. Monochromosomal cell hybrids have, however, not been available for pig gene mapping mainly owing to chromosomal instability of pig chromosomes and inter-species rearrangements. We have recently reported on the production of pig cell hybrids of an unconventional type, between pig leukocytes and a thymidine kinase (TK)-deficient cell line (MVTK super(-)) from American mink. Some of these hybrids contain a stable set of a few pig chromosomes in addition to pig Chromosome (Chr) 12, which is retained because the TK gene resides there. Thus, we were able to isolate a Chr 8-only hybrid cell line by BrdU counter selection against cells expressing TK. We here report on the construction and use of four further pig-mink hybrid cell lines with single pig Chr 2, 5, 12, or t. (1,13). Pig-mink hybrid cell lines SV27, SV29, SV35, and SV38 were characterized by G-banding. We found pig Chr 2 and 12 in 18 of 21 metaphases from SV27, pig Chr 12 and a marker chromosome in 37 of 39 metaphases from SV29, pig Chr 12 in 25 of 27 metaphases from SV35, and pig Chrs 5 and 12 in all 26 metaphases analyzed from cell line SV38. The latter cell line consisted of two cell populations: the first is presumably the progenitor and contains pig Chrs 5 and 12 on the background of a single chromosome set of MVTK super(-). The second population resulted from tetraploidization. It contains doubled pig Chrs 12 and 5 on the background of two chromosome sets of MVTK super(-). Finally, we analyzed at least 30 metaphase spreads from each of the cell lines SV27, SV29, SV35, and SV38 for the presence of rearrangements between pig and mink chromosomes by in situ hybridization with tritiated pig DNA as described earlier. No chromosomal rearrangements between pig and mink chromosomes were detected. Because of this and the presence of the same number of pig chromosomes in nearly all metaphases analyzed, we consider the four hybrid cell lines quite stable. The only pig chromosome in SV35 was Chr 12. The presence of pig Chr 12 in SV35 was confirmed by reverse painting, that is, by fluorescence in situ hybridization (FISH) to normal pig chromosome spreads by use of biotinylated inter-SINE PCR products from the hybrid cell lines.