ANALYSIS OF THE YEAST AND HUMAN GENOMES 
MaynardV. Olson, Ph.D., Investigator 
Dr. Olson's laboratory is pursuing interrelated 
studies of the yeast and human genomes, with the 
long-range goal of defining the structure and func- 
tion of the chromosomal DNA of these organisms. 
This research has three objectives: 1) to produce 
detailed physical maps of yeast and human DNA, 
which will facilitate molecular genetic studies of 
these organisms; 2) to advance the associated meth- 
odology, which will benefit research on all higher 
organisms; and 3) to study the functional behavior 
of large genes, gene clusters, and other segments of 
mammalian chromosomes. 
The Yeast Genome 
The yeast genome is ~ 1 5 million base pairs (bp) 
in size. Depending on the genetic background of the 
yeast strain, 1 or 2 million bp represent rDNA (i.e., 
they encode RNA components of the ribosomes), 
while the remaining 12.5 million bp encompass the 
rest of the nuclear genes. Dr. Olson's laboratory has 
developed detailed physical maps of this DNA that 
show the positions of known genes relative to DNA 
landmarks, such as chromosome ends and sites at 
which particular site-specific restriction endonucle- 
ases cleave the DNA. 
More than 5,300 bacteriophage X and cosmid 
clones of yeast DNA have been analyzed by com- 
puter-based techniques and used to construct con- 
tigs (sets of overlapping clones that cover a contigu- 
ous region of the genome) . These contigs now cover 
>95% of the non-rDNA component of the yeast ge- 
nome. Most of the missing DNA is at the extreme 
ends of the chromosomes, which are inaccessible to 
standard cloning techniques. Much of the effort dur- 
ing the past year has been directed toward closure of 
the contig maps, the process of analyzing and then 
eliminating gaps between contigs. This process be- 
comes progressively more difficult as the number of 
gaps decreases, since the gaps that remain are those 
that have resisted analysis by standard methods. 
Since the last annual report the number of internal 
gaps has been reduced from 69 to 12, with a corre- 
sponding increase in average contig size from 1 60 
to 411 kbp. 
The contig maps facilitate the localization of 
newly cloned yeast genes relative to previously de- 
fined genes. To aid in this process, 186 genes that 
have been mapped genetically through the analysis 
of genetic recombination during sexual crosses have 
also been localized on the physical maps. New 
genes can be localized relative to these previously 
defined genes by DNA-DNA hybridization assays that 
identify the mapped clones that contain the new 
gene. These assays take only a few hours and employ 
a set of three small nylon filters produced by Dr. 
Olson's laboratory. During the past year the number 
of laboratories using these filters has increased from 
70 to 1 50. This convenient and reliable style of gene 
mapping is becoming an early step in the character- 
ization of a yeast gene . Early mapping allows a quick 
determination of whether the gene has been studied 
previously, thereby saving much duplication of ef- 
fort in yeast molecular genetics. 
Physical Mapping of Human Chromosomes 
The average human chromosome contains 10 
times as much DNA as the entire yeast genome. Fur- 
thermore, there have been serious difficulties in 
obtaining continuous cloned coverage of human 
chromosomes in clones that are propagated in Esche- 
richia coli. Consequently, a direct extension to hu- 
man chromosomes of the methods employed to map 
the yeast genome would be unlikely to succeed. 
As an alternative strategy, Dr. Olson's laboratory 
has been pursuing applications of the yeast artificial 
chromosome (YAC) system to the physical mapping 
of human chromosomes. YAC vectors allow large 
segments of exogenous DNA to be propagated in 
yeast as linear, artificial chromosomes. During re- 
cent years. Dr. Olson's laboratory has developed 
methods for constructing large libraries of YAC 
clones containing human DNA, for identifying spe- 
cific human genes in these libraries, and for analyz- 
ing individual clones. Particularly efi'ective has 
been the use of sequence-tagged sites (STSs) as map- 
ping landmarks. STSs, which are based on random 
tracts of DNA sequence, can be recognized using the 
polymerase chain reaction (PCR). A major advan- 
tage of STS-based physical maps is that they can be 
described in a database in sufficient detail to allow 
any laboratory access to the mapped DNA with- 
out the need for supporting archives of biological 
materials. 
A method of constructing YAC contigs, which has 
been named STS-content mapping, has been devel- 
oped. In this method, overlaps between clones are 
recognized by shared STS content. Several genome 
centers involved in the international effort to map 
the human genome have adopted STS-content map- 
ping as the first-stage method of carrying out large- 
scale physical mapping. 
Attention in Dr. Olson's laboratory is now shifting 
GENETICS 241 
