A new dominant selection marker for transformation of Pichia pastoris to soraphen A resistance.
Despite the considerable progress molecular genetics of filamentous fungi has made during the past decade, there is still an urgent need for efficiently working selectable markers for fungal transformation. Using Pichia pastoris as a host, we describe the development of a new dominant selectable marker of prokaryotic origin. This system, termed sor(R), is based upon the resistance of the bacterial enzyme acetyl-CoA carboxylase (ACCase) to the macrocyclic polyketide soraphen A, a potent inhibitor of fungal ACCase produced by the myxobacterium Sorangium cellulosum. In this study, we firstly demonstrate that the integration of a single sor(R) cassette into the P. pastoris genome confers resistance to elevated concentrations of soraphen A. Furthermore, it has been shown that the versatility of this marker can be considerably increased by splitting the sor(R) cassette, especially when successive transformations are performed on the same strain. As pronounced sensitivity to soraphen A is the rule among filamentous fungi, we expect the sor(R) marker to be a widely applicable tool for fungal transformation.
Wan
Hong
H
Molecular Biology Center of University Heidelberg (ZMBH) Im Neuenheimer Feld 282, 69120 Heidelberg, Germany. hong.wan@molbio.su.se
Sjölinder
Mikael
M
Schairer
Hans Ulrich
HU
Leclerque
Andreas
A
D000103:Acetyl-CoA Carboxylase / Q000235:genetics; D000900:Anti-Bacterial Agents / Q000494:pharmacology*; D015139:Blotting, Southern; D004269:DNA, Bacterial / Q000737:chemistry / Q000235:genetics; D004271:DNA, Fungal / Q000737:chemistry / Q000235:genetics; D005819:Genetic Markers / Q000235:genetics*; D018942:Macrolides / Q000494:pharmacology*; D010843:Pichia / Q000187:drug effects* / Q000235:genetics*; D016133:Polymerase Chain Reaction; D014170:Transformation, Genetic / Q000235:genetics*
D000900:Anti-Bacterial Agents; D004269:DNA, Bacterial; D004271:DNA, Fungal; D005819:Genetic Markers; D018942:Macrolides; C086151:soraphen A; D000103:Acetyl-CoA Carboxylase