CONSTRUCTION OF A COSMID CONTIG AND OF AN ECORI RESTRICTION MAP OF YEAST CHROMOSOME-X

We report here the construction of a complete physical map of the chro mosome X of yeast Saccharomyces cerevisiae. Fragments resulting from p artial Sau3AI digestion of DNA from a diploid strain derived from S288 C were ligated to linearized pWE15, a cosmid vector with T3 and T7 pro moters. Another library, made in the cosmid vector pOU61 cos, that lac ks T3 and T7 promoters, was also used as a source of target clones. Ch romosome-X-specific clones were sorted out by hybridization with radio labelled pulse-field-gel-purified chromosome X as a probe. Then, 254 c osmids were ordered by walking from one to another by hybridization wi th end-specific T3 or T7 RNA transcripts as probes. The construction w as put to the test by hybridization with a battery of chromosome X gen e markers, that showed that the physical map and the genetic map were colinear. The validity of the contig was further strengthened by the r esults of chromosome nested fractionation with meganuclease I-Scel. An EcoRI restriction map of the contig enabled further verification and measurement of the total length of the contig, that was found to be ap proximately 700 kb in size. In addition to providing a base for the on going yeast genome sequencing project, the physical map can be used to map any sequence belonging to chromosome X.