Global Transcriptome Machinery Engineering

The Goal

Researchers Alper et al. (2006) were interested in creating a strain of yeast with improved ethanol tolerance and production.

The Experiment

gTME

The ethanol resistance and production phenotypes are regulated by many different genes in the yeast cell, and most of these genes are unknown. This fundamental lack of knowledge meant that the researchers could not rationally engineer the desired phenotype.

The researchers describe a new technique to circumvent this problem and still elicit the desired phenotype from the yeast. This method, which they have termed global transcriptome machinery engineering, is a way of using directed evolution to engineer complex phenotypes regulated by multiple genes. Unlike methods of directed evolution used for individual proteins, this method also does not require one have gene to function knowledge before they begin directed evolution.

The process is almost exactly the same as the forms of directed evolution described in previous papers, except that instead of using random mutagenesis to create a mutant library of a single gene, mutagenesis is used to create a mutant library of a selected transcription factor. gTME works under the concept that each mutant transcription factor will drastically change how genes are expressed across the entire cell once it has been transferred in vivo. The phenotypes created by these new changed expression levels in cell can then be run through selection to select for the desired phenotype (in this case, ethanol production and resistance).

The Experiment

The team began

gTME

Instead of randomizing the genes for the desired phenotype themselves, most of which are unknown, the researchers chose to create a mutant library of a known transcription factor, SPT15. The team hypothesized that small changes to this transcription factor would affect gene expression cell wide in yeast. Hypothetically,

A certain mutated transcription factor, therefore, might cause upregulation certain genes and downregulation of others to form a robust new yeast cell, which could then be found in selection tests.

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