YAC fragmentation with repetitive and single-copy sequences: detailed physical mapping of the presenilin 1 gene on chromosome 14

We constructed new LYS2 fragmentation vectors that allow direct acentric an d centric fragmentation of yeast artificial chromosomes (YACs) and selectio n of fragmented YACs in yeast strain AB1380. The fragmentation vectors were used efficiently with repetitive (e.g., Alu), low-copy (e.g., CA-repeats) and single-copy (e.g., exons) sequences. High recombination efficiencies we re obtained in fragmenting two different CEPH YACs with the Alu consensus s equence as target sequences for homologous recombination. Analysis of the a centric Alu fragmentation panel of 788H12, containing the presenilin 1 (PSE N1) gene for familial Alzheimer's disease (AD), indicated that high-resolut ion YAC fragmentation panels covering the entire parent YAC are obtained. A lso, marker content analysis of the fragmentation panel indicated that frag mented YACs were propagated stably without rearrangements. The same fragmen tation vectors were used efficiently for fragmentation of 788H12 with uniqu e sequences, i.e., exons 3 and 12 of PSEN1 and D14S77, a polymorphic CA rep eat, as target sequences. Together, our YAC fragmentation data of 788H12 pr ovided a size estimate for the coding region of PSEN1 of 60 kb and a more p recise localization of D14S77 at 25 kb upstream of PSEN1. (C) 1999 Elsevier Science B.V. All rights reserved.