Construction of a complete genomic library of Saccharomyces cerevisiae and physical mapping of chromosome XI at 3.7 kb resolution.
A consortium of European laboratories has been organized to systematically sequence the genome of Saccharomyces cerevisiae. As part of the BIOTECH program aimed at sequencing chromosomes XI and II, we have constructed a total genomic library of yeast strain FY1679 (a direct S288C derivative) into cosmid vectors pWE15 and pOU61cos. Primary clones from four independent libraries totalling 190 genome equivalents have been stored at -80 degrees C. A subset of 1939 independent clones (six genome equivalents) was hybridized using purified chromosomes XI and X as probes. A total of 147 chromosome XI-specific cosmid clones was used to construct the physical map of that chromosome. Mapping methods included a combination of classical bottom-up strategies (fingerprinting, hybridizations) and a novel top-down strategy using I-SceI chromosome fragmentation. The 147 cosmid clones form a unique contig covering the entire chromosome XI (666 kb) with the sole exceptions of the (C1-3A)n repeats of the telomeres. Colinearity of cosmid inserts with yeast DNA was directly verified. A complete EcoRI map of chromosome XI was deduced from partial overlaps of cosmids and used for the sequencing program. Comparison of this map with the genetic map shows unexpected divergences that have been solved by subsequent genetic analysis, yet underline the necessity of independent physical mapping in genome projects.
Thierry
A
A
Départment de Biologie Moléculaire (URA 1149 du CNRS and UFR927 Université P.M. Curie), Institut Pasteur, Paris, France.
Gaillon
L
L
Galibert
F
F
Dujon
B
B
D001483:Base Sequence; D002874:Chromosome Mapping; D015825:Chromosomes, Fungal; D015698:Genomic Library; D008969:Molecular Sequence Data; D012441:Saccharomyces cerevisiae / Q000235:genetics*; D016615:Telomere