Assessment of Hansenula polymorpha and Arxula adeninivorans-derived rDNA-targeting elements for the design of Arxula adeninivorans expression vectors.
Different targeting sequences derived from the Arxula adeninivorans and Hansenula polymorpha rDNA clusters were tested in A. adeninivorans integration/expression vectors. For element identification, the rDNA unit of A. adeninivorans (accession number ) was first isolated and characterized in addition to the known H. polymorpha unit. The rDNA is a cluster of some forty 7653-bp units without the 5S rDNA gene. The selected elements were integrated into a set of A. adeninivorans expression/integration vectors harbouring a TEF1 promoter - amyA ORF - PHO5 terminator sequence as reporter gene. No differences in mitotic stability, copy number and transformation frequency were observed. All transformants harboured a single copy integrated into the rDNA by a homologous recombination. In contrast, the choice of the rDNA targeting sequence was found to be of impact on productivity. Use of ETS-18S-5.8S fragments from both organisms resulted in a more than 50% increase in comparison to the use of other elements, independent of the orientation within the vector.
Steinborn
Gerhard
G
Institut für Pflanzengenetik und Kulturpflanzenforschung, Corrensstr. 3, D-06466 Gatersleben, Germany.
Gellissen
Gerd
G
Kunze
Gotthard
G
D003001:Cloning, Molecular; D004271:DNA, Fungal; D004275:DNA, Ribosomal / Q000235:genetics*; D017930:Genes, Reporter; D005822:Genetic Vectors; D010843:Pichia / Q000235:genetics*; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D004718:Saccharomycetales / Q000235:genetics* / Q000378:metabolism; D014170:Transformation, Genetic
D004271:DNA, Fungal; D004275:DNA, Ribosomal; D011994:Recombinant Proteins