title: Fast and efficient genetic transformation of oleaginous yeast Rhodosporidium toruloides by using electroporation.
abstract: Metabolic engineering of Rhodosporidium toruloides, a robust lipid and caroteinoid producer, is of great importance for oleochemicals and carotenoids production. However, the Agrobacterium-mediated gene transformation is tedious and time consuming. Here, we described a fast and efficient genetic transformation of R. toruloides using electroporation with linear DNA fragments, and the process was optimized. The results showed that 2 × 103 transformants can be obtained at 0.7 kV/μg linear DNA by using hygromycin and bleomycin as selection markers after the competent cells pretreated with 25 mM DTT and 100 mM LiAc. Our results would facilitate mutant library construction and metabolic engineering of R. toruloides for production of oleochemicals and carotenoids. We further demonstrated that all transformants arose due to illegitimate integration of transforming DNA fragments by colony PCR.
authors.affiliation: School of Biological Engineering, Dalian Polytechnic University, Dalian 116034, China.|Dalian Institute of Chemical Physics, CAS, 457 Zhongshan Road, Dalian 116023, PR China.
authors.forename: Hongdi
authors.initials: H
authors.lastname: Liu
authors.affiliation: Dalian Institute of Chemical Physics, CAS, 457 Zhongshan Road, Dalian 116023, PR China.
authors.forename: Xiang
authors.initials: X
authors.lastname: Jiao
authors.affiliation: Dalian Institute of Chemical Physics, CAS, 457 Zhongshan Road, Dalian 116023, PR China.
authors.forename: Yanan
authors.initials: Y
authors.lastname: Wang
authors.affiliation: Dalian Institute of Chemical Physics, CAS, 457 Zhongshan Road, Dalian 116023, PR China.|Dalian National Laboratory for Clean Energy, Dalian 116023, China.
authors.forename: Xiaobing
authors.initials: X
authors.lastname: Yang
authors.affiliation: Dalian Institute of Chemical Physics, CAS, 457 Zhongshan Road, Dalian 116023, PR China.
authors.forename: Wenyi
authors.initials: W
authors.lastname: Sun
authors.affiliation: School of Biological Engineering, Dalian Polytechnic University, Dalian 116034, China.
authors.forename: Jihui
authors.initials: J
authors.lastname: Wang
authors.affiliation: Dalian Institute of Chemical Physics, CAS, 457 Zhongshan Road, Dalian 116023, PR China.|Dalian National Laboratory for Clean Energy, Dalian 116023, China.
authors.forename: Sufang
authors.initials: S
authors.lastname: Zhang
authors.affiliation: Dalian Institute of Chemical Physics, CAS, 457 Zhongshan Road, Dalian 116023, PR China.|Dalian National Laboratory for Clean Energy, Dalian 116023, China.
authors.forename: Zongbao Kent
authors.initials: ZK
authors.lastname: Zhao
mesh_terms: D018274:Electroporation / Q000379:methods*; D060847:Metabolic Engineering / Q000379:methods; D012248:Rhodotorula / Q000235:genetics*; D014170:Transformation, Genetic
keywords: Rhodosporidium toruloides; electroporation; genetic transformation; linear DNA fragment; oleaginous yeast