Yeast Lab: Basics

Question 1

Yeast

Answer 1

• Single-celled fungus.
• reproduce asexually by binary fission or by the pinching of small buds off a parent cell.
• Many fungal species can grow both as yeasts and as a network of filaments; relatively few species grow only as yeasts
• Can exist as diploid or haploid
• We are using Saccharomyces cerevisiae

Question 2

Yeast Model System

Answer 2

• Model eukaryotic organism
• Single cell
• Fast growth - 90 min.
• Phenotypes of mutant genes easily seen in haploids
• Efficient transformation
• Same cellular organization as more complex eucaryotic organisms
• Organelles w/in Yeast
  
  • Nucleus
  • ER
  • Golgi
  • Mitochondria
  • Vacuole
  • Peroxisomes
  • Does not have chloroplast
• Fairly small genome: 6000 genes
• Has 16 chromosomes

Question 3

Saccharomyces cerevisiae Mating types

Answer 3

• Haploid yeast exist as different mating types: a and α
• Each type secretes a specific factor (hormone) that binds only to receptors on the other type of cell.
  
  • When exposed to each other’s mating factors, a pair of cells of opposite type change shape, grow toward each other, and fuse (mate)
  • cell cycle is paused, cell fusion and nuclear fusion occurs
  • The new diploid a/α contains all the genes of both original cells, a combination of genetic resources that provides advantages to the cell’s descendants
• Haploid cells of opposite mating type can form diploids while cells of the same mating type cannot

Question 4

genotypes of the haploid progeny of diploid cells are the result of

Answer 4

• independent assortment of chromosomes
• recombination during meiosis

Question 5

To recombine several mutant genes in one strain you can …

Answer 5

• haploid cells can be mated to form a diploid cell
• diploid can be induced to undergo meiosis (sporulation), resulting in haploid progeny with new combinations of mutations
• sporulation is induced by plating cells on a medium lacking nitrogen
  *

Question 6

vegetative propagation

Answer 6

Asexual reproduction in plants that is facilitated or induced by humans

Question 7

Agent of Fermentation

Answer 7

• Perform alchoholic fermentation: 1 glucose → 2 EhOH + 2CO₂
• xUtilize sugar and release ethanol
• xFermentation associated with yeast growth
• xFermentation failed when yeast destroyed by heat
• Zymase: active agent a protein

Question 8

Life Cycle

Answer 8

• Two forms of yeast cells can survive and grow
  
  • haploid
    
    • asexual form of the fungus
    • undergo mitosis and growth
    • under conditions of high stress will, in general, die
  • diploid.
    
    • sexual form of the fungus
    • undergo mitosis and growth
    • reproduces by mitosis when nutrients are abundant
    • When nutrient limited/stressfull conditions
      
      • cells undergo sporulation and enter meiosis
      • produce 4 haploid spores encased in a protective outer layer called the ascus
      • haploid stores can be of both mating types
      • Confinement within the ascus is thought to enforce mating between products of the same meiotic division
• life cycle
  
  • mating between haploids of opposite mating types
  • mating produces diploid cells: a/α
  • Vegetative growht of diploid cells via budding
  • Starvation induces ascus formation and meiosis
  • Meiosis generates 4 haploid ascopores encased in an ascus
  • Ascus ruptures and spores germinate
  • Haploid cells undergo vegetative growth
• rate of mitotic growth differs between haploid and diploid cells
• Under optimal conditions, yeast cells can double their population every 90 minutes
• growth rates vary between strains and environments
• Mean replicative lifespan is about 26 cell divisions
• In the wild, recessive deleterious mutations accumulate during long periods of asexual reproduction of diploids, and are purged during selfing (genome renewal)

Question 9

Quiz Question

What is complementation?

Give an example we are using in Mol Gen lab.

Answer 9

• Complementation is the restoration of the wild type phenotype in a mutant.
• The method of restoration can vary.
• One example is the restoration of the wild type phenotype of the yeast met3 mutant by the Arabidopsis MET3 cDNA in the Cloning of a Higher Plant Gene project.
• Each of the 3 yeast projects has examples of complementation.