Workshop: Yeast Genetics

Saccharomyces cerevisiae as a model organism

used to study aging, cell cycle, apoptosis, gene expression and metabolism

uses of yeast and fungi in biotechnology

fermentations, production of biopharmaceuticals, biocatalysts and recombinant proteins

doubling time of S. cerevisiae at 30°C

90 minutes

what is tetrad analysis used for?

mapping mutations and constructing strains

How is tetrad analysis carried out?

a and α strains are crossed. Meiosis forms 4 haploid progeny which are dissected and analysed

parental ditype (PD)

offspring same configuration as parent strains

nonparental ditype (NPD)

completely recombinant progeny - none of the parental genotypes are present

tetratype

offspring are one of each possible genotype/class

What does it mean if more tetrads are PD than NPD?

the observed genetic loci are linked

1 PD: 1 NPD: 4T

indicates independent assortment - no gene linkage

What happens in the tetrad if loci are linked to their respective centromeres?

proportion of T decreases

Homologous Recombination

Technique for gene knock-out or knock-in. CRISPR-Cas used for gene editing.

Yeast Two-Hybrid experiments

GAL4 domain-fused bait and prey proteins interact leading to transcription of reporter genes

Why does the Takara system (variation of hybrid experiment) have 4 different promoters?

to reduce false positives

Protein Localisation

determining location of proteins within cells using fluorescent reporter genes like GFP and immunofluorescence.

What is important for protein localisation?

position of the tag at the N or C terminal and choice of reporter gene

results of mutagenesis (random or targeted)

can lead to development of new isolates with improved productivity and/or growth

What allows us to design strategies to optimise production yields in genetic engineering?

We need to understand yeast metabolism and biosynthetic pathways

Artemisinin Semi-Synthesis

Using yeast as a host strain for heterologous expression of Artemisia annua genes in the production of artemisinin.