Jeff Young, Botanist youngbiol'wwu'edu x3638 Office: BI412

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Jeff Young, Botanistyoung_at_biol.wwu.edux3638Offi
ce BI412
Office Hours M 3 - 4 W 2 - 3 F 3 - 4
Arabidopsis thaliana Genome-based, molecular
study of plant physiology and environmental
responses.
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Omics

• Genomics the cloning and molecular
  characterization of entire genomes,
• Genome the entire complement of genetic material
  in a cell,
• Transcriptomics genomic scale gene expression
  studies,
• Proteomics the study of the proteins produced by
  the genome.

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Interactomeour focus

• Proteins interact with DNA

Proteins interact with Proteins
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Our Focus

• Reverse Genetics,
• DNA Microarrays,
• expression analysis,
• identifying cis-acting elements,
• Yeast-2-Hybrid,
• Mass Spectometry.

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Nature Reviews Genetics
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Yeasteukaryote model organism

• Eukaryote
• mitochondria,
• organelles,
• cell cycle, etc.
• Eukaryote Plus
• haploid, diploid,
• extra-chromosomal DNA.

Saccharomyces cerevisiae Bakers Yeast
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Yeast Genome Project

• Yeast Genome Project finished in 1996,
• 1.2 x 107 DNA base pairs,
• 16 chromosomes, 230 kb - 2, 352 kb,
• 6,000 Open Reading Frames (ORFs),
• Only 4 of the genes have introns,
• gt 70 of the genome is coding.

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Yeast Genome Projectvs. human genome

• 12.1 Mb Genomic DNA sequence (Human, 3,000 Mb)
• 70 coding sequence (Human, 1.8)
• Few Introns (Humans many)
• 6012 Genes (Human, 30-40,000)

About 70 of the genes found in humans, are found
in yeast.
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Known/Unknown

• 3,780 genes with some characterization
• 560 homologous with other organisms
• 1900 unknown

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Assigning Gene Function

• Biochemist enzymatic function,etc.
• Geneticist gene sequence, expression, etc.
• Cell Biologist cellular location, etc.
• - especially -
• Protein/Protein Interactions
• Protein/DNA interactions
• Etc.

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The Awesome Power of Yeast Genetics

• Homologous Recombination
• Transposons
• Life Cycle
• etc.

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Homologous Recombination

• the replacement of a gene with an exogenous gene
  through equal crossing over,

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Transposons

• Transposons whole units of DNA that have the
  ability to insert themselves into DNA molecules,
• can carry other genes.

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Hologous Recombination and Transposons

• Serve as shuttles to carry experimental DNA
  sequences into yeast,
• Regulatory sequences (promoters) drive the
  expression of,
• Reporter Genes code for enzymes that signal
  their presence in specific cells,
• Epitope Tags code for a foreign protein that
  binds to a specific antibody,
• etc.

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Yeast Life Cycle
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Reverse Genetics
Function
Classical Genetics
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Transposon Down Sidesrandom can be unsatisfying

• Insertions are essentially generated at random
• it is very difficult to mutagenize all genes
  within a genome by transposon mutagenesis alone,

• transposon-specific biases in target-site
  selection,
• for reasons not fully understood, transposons
  such as Tn3 insert non-randomly into certain
  regions of target DNA.

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Site Directed Mutagenesis

• Systematic deletion of each ORF in the genome,
• homologous recombination replaces the gene with a
  selectable marker, and a DNA barcode,
• UPTAG,
• DOWNTAG.

Whole set available1,500
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DNA Microarray

• DNA arrayed at high density on a solid substrate,
• In this experiment, DNA complementary to each ORF
  UPTAG and DOWNTAG is arrayed in an ordered
  fashion.

Fig. 8-62
http//www.bio.davidson.edu/courses/genomics/chip/
chip.html
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http//www.bio.davidson.edu/courses/genomics/chip/
chip.html
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Conditional Mutants
Conditional Mutants mutants that have observable
phenotypes under a given set of growth conditions.
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DNA Protein Interactions Interactome 1
cont. next page
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DNA Protein Interactions Interactome 1
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Wednesday

• Be ready to describe figures 4 and 5,
• Lectures online at my Course Materials Page.