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Neochromosomes: Creating transcriptional units with yeast Golden Gate assembly

Author(s): Eric Cooper

Hartwick College

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Summary:
This resource is a series of protocols using the yeast Golden Gate (yGG) method as developed by Neta Agmon and Leslie Mitchell in Jef Boeke’s lab to assemble a functional yeast gene with protein-coding and regulatory sequences.

Licensed under CC Attribution-ShareAlike 4.0 International according to these terms

Version 2.0 - published on 29 Oct 2019 doi:10.25334/3J95-S075 - cite this

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Description

This resource is a series of protocols using the yeast Golden Gate (yGG) method as developed by Neta Agmon and Leslie Mitchell in Jef Boeke’s lab to assemble a functional yeast gene with protein-coding and regulatory sequences.  First, there are protocols to assemble a single gene whose activity is easy to measure (e.g. an auxotrophic marker, antibiotic resistance gene, LacZ, etc.) just to get the system up and running in your lab.  Once you do, you can apply the methodology to any gene(s) of interest (i.e. it does not have to be a yeast gene).  You can also use a variation of this method to string together multiple genes (via VEGAS assembly) into a single plasmid to study metabolic pathways. This resource was developed as part of NSF Award #1827204, RCN-UBE: The Build-a-Genome Network.

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