Open AccessFunctional integration of multiple genes into the genome of the oleaginous yeast R hodosporidium toruloides
Author(s) -
Lin Xinping,
Wang Yanan,
Zhang Sufang,
Zhu Zhiwei,
Zhou Yongjin J.,
Yang Fan,
Sun Wenyi,
Wang Xueying,
Zhao Zongbao K.
Publication year - 2014
Publication title -
fems yeast research
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.991
H-Index - 92
eISSN - 1567-1364
pISSN - 1567-1356
DOI - 10.1111/1567-1364.12140
Subject(s) - biology , transformation (genetics) , genome , gene , agrobacterium , yeast , genetics , computational biology
The basidiomycetous yeast R hodosporidium toruloides represents an excellent producer for microbial lipids and carotenoids. However, further rational engineering of this unconventional yeast remains challenging partially because of the absence of efficient and reliable transformation method. In this study, we developed an A grobacterium ‐mediated transformation ( ATMT ) protocol for effective gene integration into the R . toruloides genome. Both haploid and diploid strains were successfully modified, and the integration was confirmed by colony PCR , Western blot analysis and genome walking. We further demonstrated that multiple genes could be integrated by consecutive ATMT , leading to engineered strains simultaneously resistant to multiple antibiotics. Our results provided a practical method for functional integration and expression of exogenous genes in R . toruloides , which should facilitate the development of genetic tools and the construction of superior strains to produce biofuel molecules and biochemicals.