title: The genome sequence of the popular hexose-transport-deficient Saccharomyces cerevisiae strain EBY.VW4000 reveals LoxP/Cre-induced translocations and gene loss.
abstract: Saccharomyces cerevisiae harbours a large group of tightly controlled hexose transporters with different characteristics. Construction and characterization of S. cerevisiae EBY.VW4000, a strain devoid of glucose import, was a milestone in hexose-transporter research. This strain has become a widely used platform for discovery and characterization of transporters from a wide range of organisms. To abolish glucose uptake, 21 genes were knocked out, involving 16 successive deletion rounds with the LoxP/Cre system. Although such intensive modifications are known to increase the risk of genome alterations, the genome of EBY.VW4000 has hitherto not been characterized. Based on a combination of whole genome sequencing, karyotyping and molecular confirmation, the present study reveals that construction of EBY.VW4000 resulted in gene losses and chromosomal rearrangements. Recombinations between the LoxP scars have led to the assembly of four neo-chromosomes, truncation of two chromosomes and loss of two subtelomeric regions. Furthermore, sporulation and spore germination are severely impaired in EBY.VW4000. Karyotyping of the EBY.VW4000 lineage retraced its current chromosomal architecture to four translocations events occurred between the 6th and the 12th rounds of deletion. The presented data facilitate further studies on EBY.VW4000 and highlight the risks of genome alterations associated with repeated use of the LoxP/Cre system.
authors: Department of Biotechnology, Delft University of Technology Julianalaan 67, 2628 BC Delft, The Netherlands.
authors: Daniel
authors: D
authors: Solis-Escalante
authors: Department of Biotechnology, Delft University of Technology Julianalaan 67, 2628 BC Delft, The Netherlands.
authors: Marcel
authors: M
authors: van den Broek
authors: Department of Biotechnology, Delft University of Technology Julianalaan 67, 2628 BC Delft, The Netherlands.
authors: Niels G A
authors: NG
authors: Kuijpers
authors: Department of Biotechnology, Delft University of Technology Julianalaan 67, 2628 BC Delft, The Netherlands Platform Green Synthetic Biology, Julianalaan 67, 2628 BC Delft, The Netherlands.
authors: Jack T
authors: JT
authors: Pronk
authors: Institute for Molecular Biosciences, Goethe-University, Max-von-Laue-Str. 9, D-60438 Frankfurt, Germany.
authors: Eckhard
authors: E
authors: Boles
authors: Department of Biotechnology, Delft University of Technology Julianalaan 67, 2628 BC Delft, The Netherlands Platform Green Synthetic Biology, Julianalaan 67, 2628 BC Delft, The Netherlands.
authors: Jean-Marc
authors: JM
authors: Daran
authors: Department of Biotechnology, Delft University of Technology Julianalaan 67, 2628 BC Delft, The Netherlands p.a.s.daran-lapujade@tudelft.nl.
authors: Pascale
authors: P
authors: Daran-Lapujade
mesh_terms: D001692:Biological Transport; D004271:DNA, Fungal / Q000737:chemistry / Q000235:genetics*; D017353:Gene Deletion; D015321:Gene Rearrangement; D016681:Genome, Fungal; D006601:Hexoses / Q000378:metabolism; D007621:Karyotyping; D060847:Metabolic Engineering; D012441:Saccharomyces cerevisiae / Q000235:genetics* / Q000378:metabolism; D017422:Sequence Analysis, DNA; D014178:Translocation, Genetic
chemical_list: D004271:DNA, Fungal; D006601:Hexoses
keywords: Cre recombinase; LoxP sites; chromosomal translocations; hexose transport; sequential deletions; yeast