Using high-molecular-weight DNA fragments from a human lymphoblastoid cell line, a pilot collection of 2500 YACs was constructed in YKK115, a recombination-deficient strain of Saccharomyces cerevisiae carrying mutations in both the rad51 and rad52 genes. Analysis of 520 clones from the current library by pulsed-field gel electrophoresis revealed more than 97% single YACs with an insert size averaging 340 kb. Fluorescent in situ hybridization (FISH) performed with 37 clones on metaphase chromosomes suggested a high proportion mapping at centromeric (7) or telomeric (4) locations. The results are consistent with the stabilization of YACs in strains disarmed in recombination functions [Kohno, K., Oshiro, T., Kishine, H., Wada, M., Takeda, H., Ihara, N., Imamoto, F., Kano, Y. and Schlessinger, D. (1997) Human YACs unstable in a rad52 single mutant strain become stable in rad51rad52 double mutant. Gene, 000, 000-000 (GENE 10429)], and further suggest that the YACs may include regions that have been difficult to clone in other strains.

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Journal Article | Research Support, Non-U.S. Gov't | Research Support, U.S. Gov't, P.H.S.

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Palmieri G, Miano MG, Casamassimi A, Lania A, Kohno K, Schlessinger D, D'Urso M, Featherstone T

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