Yeast databases and current functional
genomics studies in yeast
鄭明媛
e-mail: mingch@ym.edu.tw
Fax:(02)2826-4930
Tel: (02)2826-7045
I. Yeast databases
SGD (SGDTM) (Saccharomyces Genome Database)
SGDTM is a scientific database of the molecular biology and
genetics of the yeast Saccharomyces cerevisiae, which is commonly
known as baker's or budding yeast
http://genome-www.stanford.edu/Saccharomyces/
MIPS (Munich Information center for Protein Sequences)
The MIPS yeast genome database (MYGD) presents a
comprehensive database which summarizes the current knowledge
regarding the more than 6000 ORFs encoded by the Yeast Genome
http://www.mips.biochem.mpg.de/proj/yeast
YPD (Yeast Proteome Database) Not accessible from commercial I.P.
addresses
The Yeast Proteome Database, YPDTM, contains an up-to-date
accumulation of knowledge on all the proteins of an eukaryotic cell,
the yeast Saccharomyces cerevisiae. This yeast is the most
thoroughly studied living organism, with over 3000 of Its 6000
proteins characterized by either biochemistry or genetics or both.
The wealth of knowledge on these proteins can be applied as a model
to gain rapid insights on the cellular functions of all eukaryotic cells
from fungi to humans.
http://www.proteome.com/databases/index.html
The Virtual Library - Yeast
http://genome-www.stanford.edu/Saccharomyces/VL-yeast.html
II. Functional genomics
1. Large-scale deletion and mutational analysis
a. genetic footprinting
b. Yeast deletion project
c.Transposon-tagged disruption approach
http://www.ygac.med.yale.edu/
2. xpression analysis
a. by SAGE
b. by array-based hybridization technology
e.g. Cell Cycle Regulation
http://genome-www.stanford.edu/cellcycle/
3. the yeast proteome
a. by 2-D gel and mass-spectrometry
4. network of protein interactions - protein-linkage map
a. The Yeast Protein Interaction Map Project
http://depts.washington.edu/sfields/projects/YPLM
How the two-hybrid system works
If the two proteins interact, the reporter gene (here: HIS3) is switched on and the
diploids can grow on -His plates:
If the two proteins don't interact, the reporter gene remains inactive and the cells
can't grow on -His plates:
How to make 6000 GAL1-AD clones
For each of the ~6000 yeast
ORFs a specific primer pair
was synthesized (Research
Genetics, Alabama) and the
ORF amplified from genomic
DNA.
Each of the forward primers
had a specific common tail as
much as the reverse primers,
which had a different common tail
(20 nucleotides) .
The common tails served as
priming sites for a second
round of PCR (the “rePCR”).
The primers for this re-PCR
were 70mers with 50
nucleotides of yet another set
of common tails. Since all
primary PCR products had
already common 20mers at
their ends, one pair of 70mers
was sufficient to re-PCR all
6000 ORFs
The re-PCR products with the
common 50-mers were then used to
recombine the ORFs into twohybrid vectors in frame with the
GAL4 activation domain
Recombination cloning can be
achieved by simply transforming
linearized vector and PCR product
into yeast. The only requirement
for recombination is an overlap
between insert and vector of at
least 30-40 base pairs.
Analysis of protein-protein interaction (PathCalling)
e.g.AKR1
http://portal.curagen.com/extpc/com.curagen.portal.servlet.P
ortalYeastList
III. Cross-species comparison
a. yeast genes and human disease genes
b. ortholog vs paralog
http://www.ncbi.nlm.nih.gov/XREFdb/
IV. Yeast resources for funcional genomic studies
a. deletion strains (from Research Genetics or from ATCC)
http://www.resgen.com/products/YEASTD.php3 - ordering
Search form
http://www-deletion.stanford.edu/cgi-bin/deletion/search3.pl
b. yeast expression vectors (from Research Genetics)
YES-GS based Genestorm clones (to study individual
genes)
http://www.invitrogen.com/genestorm/index.html
GST-fusion clones (use in genome-wide manner)
http://www.resgen.com/products/YEXclns.php3
c. Yeast GeneFiltersR Microarrays
http://www.resgen.com/products/YeastGF.php3
d. Stanford microarray lab.
http://cmgm.stanford.edu/pbrown/