New families of single integration vectors and gene tagging plasmids for genetic manipulations in budding yeast.

Details

Ressource 1Download: 5_27637489_Postprint.pdf (4664.01 [Ko])
State: Public
Version: Author's accepted manuscript
Serval ID
serval:BIB_AE8913A1487C
Type
Article: article from journal or magazin.
Collection
Publications
Institution
Title
New families of single integration vectors and gene tagging plasmids for genetic manipulations in budding yeast.
Journal
Molecular Genetics and Genomics
Author(s)
Wosika V., Durandau E., Varidel C., Aymoz D., Schmitt M., Pelet S.
ISSN
1617-4623 (Electronic)
ISSN-L
1617-4623
Publication state
Published
Issued date
2016
Peer-reviewed
Oui
Volume
291
Number
6
Pages
2231-2240
Language
english
Abstract
The tractability of the budding yeast genome has provided many insights into the fundamental mechanisms regulating cellular life. With the advent of synthetic biology and single-cell measurements, novel tools are required to manipulate the yeast genome in a more controlled manner. We present, here, a new family of yeast shuttle vectors called single integration vectors (pSIV). Upon transformation in yeast, these plasmids replace the entire deficient auxotrophy marker locus by a cassette containing an exogenous marker. As shown using flow cytometry, this complete replacement results in a unique integration of the desired DNA fragment at the marker locus. In addition, a second transcriptional unit can be inserted to achieve the simultaneous integration of two constructs. The selection marker cassettes, present in the pSIV, were also used to generate a complete set of gene tagging plasmids (pGT) encompassing a large palette of fluorescent proteins, from a cyan fluorescent protein to a near-infrared tandem dimer red fluorescent protein. These tagging cassettes are orthogonal to each other thanks to the use of different TEF promoter and terminator couples, thereby avoiding marker cassette switching and favoring integration in the desired locus. In summary, we have created two sets of robust molecular tools for the precise genetic manipulation of the budding yeast.

Keywords
Genes, Reporter, Genetic Vectors/genetics, Genome, Fungal, Luminescent Proteins/genetics, Plasmids/genetics, Promoter Regions, Genetic, Saccharomycetales/genetics, Transformation, Genetic
Pubmed
Web of science
Open Access
Yes
Create date
28/09/2016 17:29
Last modification date
20/08/2019 15:18
Usage data