Worm Breeder's Gazette 11(5): 14
These abstracts should not be cited in bibliographies. Material contained herein should be treated as personal communication and should be cited as such only with the consent of the author.
In the course of the experiments we carried out in collaboration with Bob Waterston to produce a method for linking genomic sequences cloned in yeast artificial chromosomes (YACs) making use of repetitive probes (Cangiano et al. 1990, Nucl. Acids Res., 18, 5077-5081.), we came across several minor problems due to the very nature of YACs. A good example of that is what we observed on three YACs (Y39C11, Y52D1, Y43E11), we identified in a first screening as overlapping because they shared common bands with two independent repetitive probes. When we tried to reproduce these patterns with a new YAC DNA preparation, one of these YACs (Y39C11) seemed to have lost the ability to hybridize to one probe (pRcS5) and showed a different pattern with the other (pRcD1) . What had happened? We analyzed a few independent colonies from the same YAC strain (Y39C11) on a pulse field gel. The original YAC strain turned out to contain not one but two YACs, one of which in fact overlapped the other YACs analyzed. The two YACs were present in some colonies of the strain together but more often one or the other was lost, furthermore when they were present together often the longer one was also partially deleted and additional fainter shorter bands were present when the pulse field gel was hybridized with a suitable cosmid probe (ZK384). We may explain these events considering that the yeast host is under selective pressure to contain a copy of URA3 yeast gene carried by the YAC, but a single copy per cell is sufficient to restore prototrophy, so if two YACs are present together one or the other can be easily lost and/or undergo various rearrangements. The occurrence of these rearrangements can be explained in several ways: i) high frequency of recombination in yeast, ii) the occurrence of segregator sequences ( SEG sequences described by Stinchcomb et al. 1985 Proc. Natl. Acad. Sci. USA vol. 82, 4167-4171) might very well lead to dicentric YACs and eventually to their destruction, iii) many eukaryotic regulatory sequences are common to different species and we know that C. elegans DNA may actively interact with nuclear machineries in yeast (La Volpe et al. (1988) Nucl. Acids Res. 16, 8213-8231); some of these sequences may be selected against leading to rearranged chromosomes. We should all be as cautious is possible handling YACs and in general suspicious of the organization of cloned regions of YAC origin.